Biochemistry and cell biology introduction

2
Biochemistry and cell biology

Patricia Revest
Introduction
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Basic chemistry
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Chemical bonds
Basic chemical reactions
Inorganic molecules and ions
Biological compounds
Simple organic compounds
Macromolecules
Proteins
Carbohydrates
Lipids
Nucleic acids
Basic cell structure

Histology
Organelles, structure and function
Intracellular and extracellular fluids
Protein synthesis and processing

Genes and the genome
The genetic code
Transcription
Translation
Processing pathways for proteins
Protein production in the cytosol
Membrane structure and functions

Fluid mosaic model of membranes
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INTRODUCTION
The human body is mainly water plus a wide variety of bio
logically active chemicals which subserve all the functions of
the body. Biochemistry describes how these molecules are
made and the interactions between them at molecular level.
Cell biology then goes on to describe how the biochemicals
are organised into cells and cellular components, which then
form the tissues of the body. Normal processes within the
body are called physiological, whereas processes that
cause disease are called pathological.
Metabolism
All the processes of replication, growth, repair and so on that
are vital to the survival of a cell require energy. Much of the
economy of any cell is taken over by the chemistry required,
firstly, to generate that energy, and, secondly, to harness it
to drive crucial processes or to build molecules needed by
the cell to maintain itself. All this chemistry is termed metabolism. We can think of cells as being extremely complex
chemical machines. At any time a cell may have many thou
sands of individual chemical reactions going on simultan
eously, each a part of a metabolic pathway that may result
in the breakdown of larger molecules, or the assembly of
ch02.indd 15
Specialised plasma membrane structures

Cellular transport processes
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Cell-to-cell communication
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Types of signalling
Ligands and receptors
Cell recognition
Cell adhesion molecules
Cell division and DNA replication

Stem cells
Phases of the cell cycle
Mitosis
Meiosis
DNA replication
Cellular ageing and cell death
Key metabolic pathways

Glycolysis
The metabolism of fatty acids
TCA cycle
Amino acid metabolism
Electron transport chain and oxidative
phosphorylation
Tissues and organs

Origins of cells
Basic tissue types
Organs and systems
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large molecules from smaller ones, or simply the conversion

of one molecule into another. All of these pathways need to
be controlled so that molecules are produced in the right
place in the right amounts.
Catabolism and anabolism

In general, metabolism that involves degrading complex mol
ecules to simpler ones, called catabolism, is the route by
which chemical energy is generated. By contrast, the metab
olism that results in the synthesis of more complex molecules
from simpler components, anabolism, has a net requirement
for chemical energy. It is these synthetic metabolic pathways
that produce the components needed for cell growth, repair
and replication.
Cells
The cell is the basic unit of living organisms. All living things
consist of cells and although there are many organisms
which exist as single cells, such as bacteria and amoeba, the
largest living organisms on Earth, such as the great blue
whale, also consist of aggregates of the same basic cellular
units, although in rather large numbers. An adult human has
somewhere in the region of 1015 cells.
All living organisms are made up of one of two types of
cell: prokaryotes and eukaryotes.
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16 Biochemistry and cell biology

Table 2.1 Eukaryotic kingdoms
Kingdom
Examples
Examples of pathogens
Protists
Protozoans, algae, slime moulds
Plasmodium spp. (malaria)
Fungi
Yeasts, moulds, mushrooms
Candida albicans (candidiasis or thrush)
Plants
Mosses, ferns, conifers, flowering plants
None, although many plants are poisonous to humans
Animals
Sponges, corals, arthropods, amphibians, reptiles, birds, mammals
Taenia solium (tapeworm)
Table 2.2 Chemical elements of the human body

Element (atomic symbol)
Abundance (% wet weight)
Major elements
Oxygen (O)
Carbon (C)
Hydrogen (H)
Nitrogen (N)
Total
65.0
18.5
9.5
3.3
96.3
Minor elements (in order of abundance)
Calcium (Ca), phosphorus (P), potassium (K), sulphur (S), sodium (Na),
chlorine (Cl), magnesium (Mg)
Total
3.7
Trace elements (in alphabetical order)
Aluminium (Al), boron (B), chromium (Cr), cobalt (Co), copper (Cu),
fluorine (F), iron (Fe), manganese (Mn), molybdenum (Mo), selenium (Se),
silicon (Si), tin (Sn), vanadium (V), zinc (Zn)
Prokaryotes
Two of the major kingdoms into which organisms are classi
fied are the Eubacteria and the Archaebacteria. These are all
prokaryotes. Prokaryotic cells lack a nucleus and other inter
nal structures. All prokaryotic organisms are single celled;
they are extremely numerous and can be found in almost all
environments, including some which seem too hostile to
support living things. The earliest organisms on Earth were
prokaryotes, and they were the only living things for more
than 2 billion years. From the point of view of medicine, the
bacteria are important in that they cause many of the infec
tious diseases.
Many chemicals and chemical processes which occur in
the human body are similar to those found in many other
organisms, including prokaryotes. However, there are also
significant differences and sometimes these can be exploited,
for example in the use of antibiotics that can kill bacterial
cells that have invaded the body, without killing the host, by
exploiting differences in their biochemistry.
are between 1m and 10m and many eukaryotic cells are

10100mm.
BASIC CHEMISTRY
Chemical composition of the human body
The chemicals which make up the human body are mainly
compounds consisting of combinations of carbon, hydro
gen and oxygen atoms (Table 2.2) with small quantities of
other elements. These are known as organic compounds
because they contain carbon. Together with nitrogen,
carbon, hydrogen and oxygen make up 96.3% of the body
weight. The remaining 3.7% is made up of seven minor
elements. As well as these major and minor elements there
are a number of trace elements which are required in very
small amounts (less than 0.01%). Many of these are needed
as special components of proteins called enzymes and are
required for their activation (see Regulation of enzyme
activity).
Eukaryotes
The other four kingdoms are made up of organisms with
eukaryotic cells (Table 2.1). These cells have a nucleus and
other internal structures which allow compartmentation of
chemical processes within a single cell. The Protista contains
both unicellular and multicellular organisms and it is thought
that the organisms in the other three kingdoms evolved from
ancient protists.
Viruses
Although viruses are not cells and as such are not strictly
living organisms, they are dependent on the hosts cells to
reproduce, i.e. they are parasites. The effects of this parasit
ism are the symptoms of many viral infections. However, as
with bacteria, differences in chemical processes used by
viruses can be exploited to produce antiviral drugs.
Cells and viruses exist on different scales. Viruses are
very small (6080nm), which is smaller than some of the
intracellular components of eukaryotic cells. Most bacteria
ch02.indd 16
CHEMICAL BONDS
In order for atoms to combine to form molecules, chemical
bonds must be formed between the individual atoms
(Fig. 2.1). There are three main types of chemical bonds:

Ionic
Covalent
Hydrogen.

The strength of the different types of bond varies

enormously (Table 2.3). Stronger bonds are more stable but,
often, large numbers of weaker bonds stabilise molecular
interactions.
Atomic structure
In order to understand how chemical bonds are formed it is
necessary to know something about the structure of atoms.
All atoms of a particular element have a nucleus of protons
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Basic chemistry 17
A
Ionic bonds
Cl
Na+
Ionic bonds
Covalent bonds
When dissolved in water many atoms are able to form

charged particles called ions by either losing or gaining elec
trons from their outer shell:
H
H
Atoms which gain an electron become negatively

charged anions.
Atoms which lose an electron become positively
charged cations.
H
C
Hydrogen bonds
+
H
+
H
O
Fig.
+
H
+
H
+
H
These ions will be attracted to each other by the opposite

electrical charges, resulting in the formation of ionic bonds.
The tendency of an atom to lose or gain electrons depends
on the number of electrons already present.
+
H

O H
H +
2.1 Different types of chemical bond.
Table 2.3 Energy of some typical chemical bonds in

biological systems
Type of bond
Energy (kJ/mol)
Ionic
12.629.3
Covalent (single)
210460
Covalent (double)
500710
Covalent (triple)
815
Hydrogen
4.28.4
Van der Waals interaction
4.2
and neutrons (except ordinary hydrogen which has a single

proton as a nucleus) surrounded by a cloud of electrons. The
total number of protons in an element gives its atomic
number, and the total of protons and neutrons (which may
vary between different forms or isotopes of an element)
gives the atomic mass. Carbon has six protons and an
atomic number of 6, but it can have six, seven or eight neu
trons. Its atomic mass is thus the average of the mass of the
different isotopes: 12.01. This reflects the much greater
abundance of carbon-12 (where 12 is the atomic mass) than
the other isotopes, carbon-13 and carbon-14.
Atomic shells
The electrons which surround the nucleus are found in fixed
energy levels called shells and there are strict rules about
how many electrons can occupy each shell:

The innermost shell can contain a maximum of two

electrons
The next shell can contain eight electrons
The third shell can hold up to 18.
Atoms are at their most stable when their outermost shell
is either full or contains eight electrons. The most stable
ch02.indd 17
elements which exist are the noble gases such as helium or

neon, which, with two and 10 electrons, respectively, have
full outer shells. These atoms are chemically inert, their full
outer shells rendering them non-reactive, and they rarely
form compounds with other atoms. Atoms with unfilled outer
shells are more reactive.
Sodium chloride: an example of an ionic bond

Common salt, or sodium chloride, is a compound formed by
the combination of sodium and chlorine atoms. Sodium has
11 electrons: both its inner two shells are full and it has a
single electron in its outer shell. Chlorine has 17 electrons,
again with full inner shells but with seven electrons in its outer
shell. In water, sodium (Na) atoms lose a single electron to
become sodium (Na+) cations with a single positive charge.
Chlorine (Cl) atoms gain the electron lost by the sodium to
become chloride (Cl) anions with a single negative charge.
When sodium chloride solution is dehydrated, then the ions
form a lattice of interlocking anions and cations held together
by the ionic bonds between the ions to give white salt crys
tals (Fig. 2.1A). The resulting sodium chloride has completely
different properties from its component parts. Sodium is a
highly reactive metal and chlorine is a poisonous green gas,
but sodium chloride is the main ionic constituent of seawater,
where life was thought to have evolved, and forms the major
ions in the extracellular fluid which bathe all the cells in the
body.
Ionic bonds are frequently formed between molecules
such as potassium (K) or calcium (Ca), which have either one
or two electrons in their outer shell forming K+ and Ca2+,
respectively and fluorine or iodine, which like chlorine
require only one electron to fill their outer shell.
Hydrogen ions
Hydrogen ions are formed by losing its single electron to
form a cation (H+). This ion is stable because its nucleus with
its lone proton has an empty electron shell. However, hydro
gen can also form an anion (H) with more reactive metals,
called a hydride, with two electrons in its single shell. Water
forms a small number of ions when it dissociates into H+ and
OH (hydroxyl) ions. This is depicted as H2O H+ + OH and
is reversible, as indicated by the double arrows. These ions
are important in many chemical reactions.
Covalent bonds
It is not necessary for an atom to completely lose or gain an
electron in order to participate in chemical bonding. Atoms
can share their electrons with other atoms to fill their outer
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electron shells part of the time. Up to three pairs of electrons
can be shared, producing increasingly strong links known as
covalent bonds. These bonds can form between atoms of
the same element, as in the formation of O2, where two
oxygen atoms, each with six electrons in the second shell,
share two pairs of electrons forming a double bond (O=O).
Two hydrogen atoms form molecules of H2 by sharing their
electrons with a single bond (HH). When a single atom of
oxygen shares electrons with two atoms of hydrogen the
result is the most abundant biological molecule of them all,
water or H2O.
Carbon bonds
Carbon can form a wide variety of covalent bonds with
other atoms by sharing four of its electrons to form pairs
(Fig. 2.1B). When a single carbon atom combines with four
hydrogen atoms the simplest organic molecule, methane
(CH4), is formed. However, carbon can form long chains of
atoms both with other carbon atoms and with other atoms
to form complex biochemicals.
Stereoisomers
The four covalent bonds around a carbon atom are arranged
in a tetrahedral pattern. When the four chemical groups are
different this means that the molecule can exist as two mirror
images, which cannot be superimposed. The asymmetric
carbon atom is said to be chiral and the two molecules are
called stereoisomers. Stereoisomers rotate polarised light
in either a clockwise, or dextrorotatory (+), direction or in an
anticlockwise, or levorotatory (), direction.
Stereoisomers can only be converted, one to the other,
by breaking and re-forming the covalent bonds. A molecule
with one chiral centre has two stereoisomers, and one with
n chiral centres has 2n possible stereoisomers. This means
that many organic molecules with identical formulae and
chemical groups can exist in different forms depending on
the number of asymmetrical carbon atoms. This is particu
larly important in the structures of macromolecules such as
proteins and carbohydrates, in which the building blocks
consist of specific stereoisomers.
Carboncarbon double bonds

Carbon atoms can also share two pairs of electrons to form
carboncarbon double bonds. These bonds are planar and
rigid and do not rotate. When they occur in carbon chains
they produce either cis or trans isomers depending on
whether the next carbon atoms are:

On the same side of the double bond (cis)

On opposite sides (trans).
This isomerism is important in the structures of fatty

acids, which are components of many lipids. Most important
dietary fatty acids are in the cis form but commercially pro
cessed fats in foods such as some margarines and peanut
butter contain trans forms, which are known to raise blood
cholesterol. Recent nutritional guidelines recommend that
consumption of these types of fatty acids be limited.
Polar and non-polar covalent bonds

In covalent bonds between atoms, the electrons are not
always equally shared between the atoms. When the elec
trons are more attracted to one of the atomic nuclei, then the
bond has two poles of charge called a dipole and the bond
is called a polar covalent bond. This occurs in H2O, where
ch02.indd 18
the electrons are closer to the oxygen atom than the hydro
gen atoms. The oxygen then has a partial negative charge
() and the hydrogen atoms partial positive charges (+).
These partial charges affect the way these molecules react,
and are particularly important in the way water molecules
interact. In compounds such as methane, where the elec
trons are shared equally, the bonds are called non-polar
covalent bonds.
Hydrogen bonds
The partial positive charges caused by the unequal sharing
of electrons by hydrogen atoms are attracted to the partial
negative charges on atoms such as oxygen and nitrogen.
This forms a weak bond called a hydrogen bond. Although
this type of bond is only about 1/20 as strong as a covalent
bond, the presence of a large number of hydrogen bonds
can produce a significant cohesive force. Many large mo
lecules contain numerous hydrogen bonds between different
parts of the molecule and these help to maintain the
structure.
Hydrogen bonds can be disrupted easily, by heat and by
excess acidity, causing a molecule to lose its shape and
possibly its function. This is shown clearly in the formation
of ice crystals where the regular lattice of the frozen water
molecules is held together by the hydrogen bonds between
the molecules (see Fig. 2.1c). When ice is heated, the hydro
gen bonds are disrupted and ice becomes liquid water.
Other molecular interactions

Two types of interaction between non-polar molecules
produce attractive forces between molecules:

Van der Waals interactions

Hydrophobic interactions.
Van der Waals interactions happen when transient

dipoles occur in the electron clouds of uncharged molecules
in close proximity. The resultant force depends on the dis
tance between the molecules, as these interactions can only
occur when the atoms are 0.30.4nm apart.
Van der Waals interactions are weaker than hydrogen
bonding and are very dependent on the precise molecular
shapes of the interacting molecules. However, two interlock
ing molecules can form very many of these contacts and,
because of this, van der Waals interactions are particularly
important in specific interactions between biological mol
ecules, such as enzymes and their substrates.
Hydrophobic interactions take place between non-polar
molecules, which do not contain either ions or dipoles and
so do not interact with water molecules and are almost
completely insoluble in aqueous solution. These molecules
will aggregate together to exclude water. Such hydrophobic
interactions are due to the fact that the insertion of a nonpolar molecule into water requires energy because it involves
distorting the interactions between the polar water mol
ecules. Thus the most stable conformation of non-polar mol
ecules is to exclude water and to form structures that have
the least possible surface area exposed to water.
This exclusion of water by hydrophobic forces is the basis
of cell membrane structure. Cell membranes are largely
made up of molecules that are polar at one end, where they
interact with water, attached to long, non-polar carbon and
hydrogen chains which aggregate together and do not react
with water.
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Basic chemistry 19
BASIC CHEMICAL REACTIONS
In order for molecules to be formed, chemical bonds must
be made or broken between the constituent atoms. There are
three basic types of chemical reaction:
Synthetic or combination reactions in which bonds
form between two molecules or atoms to form a larger
molecule, e.g. X + Y XY.
Decomposition reactions where a molecule is broken
down into smaller parts, e.g. XY X + Y.
Exchange reactions where atoms are exchanged
between molecules, e.g. XY + AB XA + YB.

Some reactions are thermodynamically irreversible; that

is they will only proceed in a single direction. However, many
reactions can be reversed, although sometimes this may
require special conditions.
Oxidationreduction (redox) reactions

A commonly occurring biochemical reaction is the
oxidationreduction or redox reaction. In this reaction one
molecule loses electron(s) in a process called oxidation, with
the lost electron(s) being transferred to another molecule in
a process called reduction. The name redox derives from
the fact that, in biological systems, the molecule which often
receives the transferred electron is oxygen.
Many redox reactions also involve the loss of two hydro
gen atoms in the form of a hydrogen ion (H+) and a hydride
ion (H). Oxidationreduction reactions allow energy, in the
form of chemical bonds, to be transferred from the molecule
that is oxidised to the molecule that is reduced. The process
is best thought of as two tightly linked processes in which
there is a reduction in the potential energy of the oxidised
molecule, whereas the potential energy of the reduced mol
ecule is increased.
For example, one of the basic reactions used to provide
energy to cells by the metabolism of glucose is summarised
as:
C6H12 O6 + 6O2 6CO2 + 6H2 O
couple exergonic and endergonic reactions to transfer energy

between molecules.
Activation energy
The energy initially required to break the chemical bonds in
the reactants is the activation energy. In order to react,
molecules and atoms must collide with sufficient kinetic
energy to overcome the repulsive force between their elec
tron clouds, and this kinetic energy can be increased by
raising the temperature. Increasing the concentration of the
reactants also increases the chance of favourable collisions.
Some reactions also require energy input to alter certain
properties of the covalent bonds and others may require the
excitation of electrons from inner electron shells into the
outermost shell before the reaction can occur.
Catalysts
Special transport systems and mechanisms of compart
mentation within cells can help to increase the effective
concentration of a compound. However, large increases in
temperature are undesirable in biological systems and many
biochemical reactions would not occur without the presence
of a catalyst. Reactions between molecules occur more
readily if the reactants collide in the correct orientation.
Catalysts act to lower the activation energy of the reaction
by helping the reactant molecules to collide in a more favour
able direction. The catalysts themselves are unchanged by
the process so they can be used again and again. Biological
catalysts called enzymes allow a large number of chemical
reactions to occur at an appropriate rate and under
normal physiological conditions, such as at normal body
temperature. (see Enzymes, below)
INORGANIC MOLECULES AND IONS

We tend to think of life as being carbon-based. However, as
well as the large number of carbon-containing organic mol
ecules, there are large numbers of molecules which do not
contain carbon but are also essential for life. These include
water and many salts, acids and bases.
glucose + oxygen carbon dioxide + water

This is an oxidationreduction reaction: glucose is oxidised
to carbon dioxide, losing hydrogen, and oxygen accepts the
hydrogen and is reduced to water. However, in cellular
metabolism it does not occur in a single step. In a series of
linked reactions, the energy liberated in the oxidation of the
glucose is transferred to other molecules which can then be
used to drive other reactions.
The breaking of chemical bonds requires energy input,

whereas the formation of new bonds releases energy.
Thus chemical reactions produce:
Water makes up about 60% of the human body by weight.

The large amount of water which is present in and around
the cells has a number of functions. The most important of
these is as a solvent for all the molecules, the solutes, which
need to be in solution in order to participate in chemical
reactions. Water is also an important substrate in many of
these reactions. Many chemicals are transported around the
body in solution and, without water, the gases oxygen and
carbon dioxide could not be moved between the lungs and
the blood.
A 70kg human has approximately 42L of total body
water divided into two main compartments:
Energy in chemical reactions
Either a net release of energy (exergonic reactions)

Or a net absorption of energy (endergonic reactions).
In exergonic reactions the energy required to break exist

ent bonds is less than that released by the newly formed
bonds. The products have a lower potential energy than the
reactants. The reverse occurs in endergonic reactions, where
the products have a higher potential energy than the react
ants. Many biochemical reactions (e.g. the redox reactions)
ch02.indd 19
Water
Intracellular fluid contained within cells: about 28L

(approximately 40% of body weight).
Extracellular fluid: about 14L (approximately 20%
of body weight), divided between the interstitial fluid
of about 10.5L and the blood plasma volume of about
3.5L.
Water acts as a lubricant, preventing solid structures
rubbing against one another and causing damage. It also
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cushions the brain within the solid skull, reducing the pres
sure applied by the brains own weight to the base of the
brain. This cushioning prevents some of the damage which
could occur due to sudden movements of the head banging
the brain against the inside of the skull (see Ch. 8).
The high thermal capacity of water also allows it to be
used to dissipate heat easily from areas of high temperature,
preventing cell and tissue damage, and the large amount of
heat required to evaporate water released as sweat is a very
efficient cooling mechanism.
Table 2.4 Common functional groups and classes

of compounds (R represents the rest of
the molecule)
Group
Formula
Class
Hydroxyl
ROH
Alcohols
Aldehyde
RCOH
Aldehydes
Carbonyl
RCOR
Ketones
Carboxylate
RCOOH
Carboxylic acids
Ester
RCOOR
Esters
Amino
RNH2
Amines
Imino
RNH
Imines
Acids, bases and salts
Amide
RCONH2
Amides
Phosphate
RPO4
Organic phosphates
Acids, bases and salts are organic or inorganic compounds

which, when dissociated in water, form ions:
Sulphydryl
RSH
Thiols
Acids form hydrogen ions (H+) and anions
Bases form hydroxyl ions (OH ) and cations

Salts form anions and cations.

Acids and bases are crucially important in the control of

the acidity (pH) of the body, which is maintained within very
strict limits close to the neutral pH of 7 (see Ch. 1).
The salts sodium chloride (NaCl) and potassium
chloride (KCl) are the main ionic constituents of the fluids
outside and inside cells, respectively. Sodium and potassium
ions are essential for the transmission of nerve impulses,
and calcium is required for muscle contraction. However,
the commonest salts present in the body are the calcium
phosphates which make up the bones and teeth (see
Ch. 9).
Table 2.5 Macromolecules

Macromolecule
Monomer
Major functions
Proteins (15%)*
Amino acids
Structural material,
biological catalysts
Carbohydrates (2%)*
Sugars
Energy source, cell

surface markers
Nucleic acids
Nucleotides
Genetic information
(DNA and RNA)
Lipids (20%)*
(Many lipids are

triacylglycerols)
Energy storage, cell

membranes
*Note the percentages quoted are very variable due to the difference
in fat and protein content in different individuals. The values shown
here are a guide to their relative abundance.
MACROMOLECULES
BIOLOGICAL COMPOUNDS
SIMPLE ORGANIC COMPOUNDS
The simplest types of organic molecules are the hydrocarbons, which are composed of carbon and hydrogen. They
are divided into two groups:

Those that do not contain double bonds and are called

saturated
Those containing double bonds and are called
unsaturated.
Hydrocarbons are non-polar and hydrophobic. Long
hydrocarbon chains form the fats that make up cell mem
branes and form dense energy stores. Another distinctive
type of hydrocarbon is produced when carbon atoms form a
ring structure with six carbon and six hydrogen atoms, known
as benzene. These ring structures, called aromatic rings, may
also contain nitrogen.
Common chemical groups

Apart from hydrocarbons, all of the other biochemicals in the
body consist of molecules in which hydrogen has been
replaced by other atoms or groups of atoms. Potentially,
although there is a huge range of possible groupings of
atoms that can be formed, there are a limited number of
distinctive functional groups found on biological molecules
(Table 2.4). The reactivity of these groups determines the
roles played by the molecules that contain them and deter
mine their interactions with other molecules.
ch02.indd 20
Although a large number of small molecules are involved in

biochemical processes, it is only in the formation of large
molecules that the complexity of life becomes evident. There
are three major groups of macromolecules present in the
body. Each of these groups is formed from a number of
related small molecules, monomers, which can be linked
together by covalent bonds into polymers. The three groups
of macromolecules are:

Proteins
Carbohydrates
Nucleic acids.

A fourth group of molecules, which are not macromol

ecules in the same sense, are the lipids. These form macro
molecular complexes by hydrophobic bonding.
Together, these four groups of molecules make up
most of the remaining 40% of the human body, after water
(Table 2.5). Although each of these groups has a particular
type of building block, they often contain other molecules in
varying amounts. For example, proteins often have carbohy
drates covalently linked to them.
PROTEINS
Proteins have an enormous diversity of roles in the body and
have the most complex structures of all of the macromol
ecules. Although all proteins are made in a similar manner
and formed from similar building blocks, they have a wide
variety of structures and functions, some examples of which
are described below.
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Biological compounds 21
Hydrogen
H
Table 2.6 Amino acids found in human proteins
H
O
Amino
group H
N
H
R
Side chain
H
H
R
N
Carboxyl
group
O
C
Classification
Amino acid
Abbreviations
Non-polar aliphatic
Glycine
Alanine
Valine
Leucine
Isoleucine
Proline
Gly
Ala
Val
Leu
Ile
Pro
G
A
V
L
I
P
Non-polar aromatic
Phenylalanine
Tyrosine
Tryptophan
Phe
Tyr
Trp
F
Y
W
Polar uncharged
Serine
Threonine
Asparagine
Glutamine
Ser
Thr
Asn
Gln
S
T
N
Q
Polar negatively charged
Glutamic acid
Aspartic acid
Glu
Asp
E
D
Polar positively charged
Lysine
Arginine
Histidine
Lys
Arg
His
K
R
H
Sulphur-containing
Cysteine
Methionine
Cys
Met
C
M
Fig. 2.2 Generalised amino acid with ionised NH3+ and
COO groups in its two stereoisomers. The two

molecules are mirror images of each other and cannot be
superimposed.
Amino acids
Amino acids are the building blocks from which proteins are
made. They also have other roles as neurotransmitters, as a
source of energy, and as key intermediates in the mainten
ance of nitrogen balance.
Amino acid structure

Amino acids are a group of molecules with a common struc
ture, which consists of a central carbon atom, attached to
which are four different groups:

A hydrogen atom (H)

An amino group (NH2)
A carboxyl group (COOH)
A side chain (R), which varies between the different
amino acids.

At the normal pH of body fluids the amino and carboxyl

groups of amino acids are ionised to NH3+ and COO, respec
tively (Fig. 2.2). This produces a molecule with positive and
negative charges at opposite ends called a zwitterion.
However, when amino acids bond together to form proteins,
these charged groups combine. Thus it is only the side chains
(and the terminal groups) that remain charged in proteins.
A wide range of amino acids are found in living things,
but only 20 of these are made into human proteins (Table
2.6). The side chains vary in size and complexity, from a
single hydrogen atom in glycine to the large side chains of
tryptophan and arginine (Fig. 2.3). Each amino acid has a
three-letter and a single-letter abbreviation.
Essential amino acids

There is a continual turnover of proteins in the body. This
varies from minutes, in the case of some enzymes involved
in metabolic control, to months, in the case of the main
structural protein, collagen. Although some of the amino
acids are reused many are metabolised, which means that
even adults (who are not growing) require a daily dietary
intake of protein. Eight of the 20 amino acids found in human
proteins are called essential amino acids and must be
included as components of the diet because they cannot be
manufactured from metabolic intermediates (see Ch. 16).
Dietary protein must be broken down in the gut to its con
stituent amino acids, which are then absorbed into the
blood.
ch02.indd 21
Enantiomers
The central carbon atom is called the -carbon. In all the
amino acids except glycine, the presence of four different
groups around the -carbon means that each amino acid can
exist in two stereoisomers. These pairs of amino acids are
called enantiomers and are named d (dextro = right) and
l (laevo left). With very few exceptions, only l-amino acids
are incorporated into proteins. However, d-amino acids are
important constituents of the cell walls of bacteria, and as
such are targets for antibiotics such as penicillin. One of
the 20 amino acids, proline, is not actually an -amino acid
but an -imino acid with an NH group instead of the
amino (NH2) group. This has important structural conse
quences for proteins that contain this particular amino acid
(see below).
Classification of amino acids

Amino acids are classified into groups according to the
chemical properties of their side chains, R. For example,
amino acids with polar side chains, and particularly charged
ones, will form hydrogen bonds with water and will thus tend
to be found on the parts of the protein exposed to the
aqueous environment. Non-polar amino acids will be hydrophobic and will be found in parts of the protein not exposed
to water. Amino acids are classified as follows:
Non-polar aliphatic: these have side chains which
consist of carbon and hydrogen atoms with no
double bonds. These saturated hydrocarbons are
hydrophobic.
Non-polar aromatic: these side chains all contain
aromatic rings. They are generally hydrophobic except
for tyrosine, which has a hydroxyl group that can be
found on the hydrophilic part of proteins.
Polar uncharged: these neutral amino acids have polar
hydroxyl or amide groups. The hydroxyl groups on
serine and threonine, like tyrosine, are often on the
hydrophilic surface of proteins where they are often
modified to change the activity of the protein by
phosphorylation.
Polar negatively charged: these acidic amino acids have
carboxyl groups that are negatively charged at pH 7.

2/26/2009 6:48:26 PM

Non-polar aliphatic
CH2
CH3
CH
H
CH3
Alanine
CH3
CH
CH2
CH3
CH3
Glycine
CH
Valine
CH3
CH3
Isoleucine
Leucine
Non-polar aromatic
CH2
CH2
Phenylalanine
CH2
OH
N
H
Tryptophan
Tyrosine
Polar uncharged
CH2
OH
CH
CH2
OH
CONH2
CH2
CH2
CONH2
CH3
Serine
Threonine
Asparagine
Glutamine
Polar negatively charged

CH2
CH2
COOH
CH2
Glutamic acid
COOH
Aspartic acid
Polar positively charged

CH2
CH2
CH2
CH2
NH2
CH2
CH2
CH2
NH
C
NH
Lysine
Arginine
NH2
CH2
N
NH
Histidine
Sulphur-containing
CH2
SH
Cysteine
CH2
CH2
CH3
Methionine
Imino
Fig.
COOH
Proline
2.3 Structures of the 20 common amino acids. This diagram shows only the R group of each amino acid.
Polar positively charged: lysine and arginine are both

positively charged at pH 7. However, the side chain of
histidine is mainly charged at a pH of 6.8 and mainly
uncharged at a pH of 7.8. Variations in pH can therefore
influence whether histidine residues are charged or not.
Sulphur-containing: adjacent cysteine residues on
protein chains can form bonds between their sulphur
atoms called disulphide bonds. These are important in
maintaining the structure of the protein and in forming
links between separate protein chains.

Peptide bonds
Amino acids are linked together to make proteins by the for
mation of bonds between the carboxyl group of one amino
acid and the amino group of the other, with the elimination
of a water molecule. The reaction leaves an amide bond
called a peptide bond.
ch02.indd 22
H3N CHR COO- + +H3N CHR COO-

+
H3N CHR COHN CHR COO- + H2 O
This reaction can be repeated, adding more amino acids

to form a chain of amino acid residues with a carboxyl group
at one end, the C-terminal, and an amino group at the other
end, the N-terminal. Molecules containing two amino acid
residues are called dipeptides, those with three, tripeptides.
Usually peptides with more than 50 residues are called proteins, or polypeptides, and they can be made up of thou
sands of amino acids. Amino acid residues in a protein are
numbered from the N-terminal end, which is the end from
which protein synthesis starts.
The single covalent peptide bond has some of the rigidity
of a double bond because electrons are unevenly shared
between the COHN atoms (Fig. 2.4). This means that rotation
around the CN linkage is limited. However, rotation can still
occur around the -carbon atom and this rotation affects the
way that the chains can fold.
2/26/2009 6:48:27 PM
R
C
H
N
O
R
H
R
Peptide
Peptide
bond
bond
Fig. 2.4 The peptide bond. The peptide bond is
relatively rigid but rotation can occur around the -carbon.
Protein structure
Although proteins can be thought of as a linear sequence of
amino acids, this does not convey the fact that, in order to
function properly, each protein chain must be formed into
the appropriate three-dimensional shape. There are four
levels of structure that are used to describe proteins.
Information box 2.1 Sickle cell anaemia
Changes in a single amino acid in a protein chain can have

extreme consequences in the function of that protein. In
sickle cell anaemia, the haemoglobin protein of red blood
cells has a single change in one of the 146 amino acids
which make up the -globin chain. This is due to a
mutation in the DNA which changes a single base from A to
T, resulting in a glutamate (glutamic acid) residue at position
6 in the normal protein being replaced by a valine. This
substitution of a negatively charged side chain by a nonpolar hydrophobic one allows the proteins to form rigid
rods under low oxygen conditions, changing the shape of
the cells and making them inflexible and unable to travel
through small blood vessels as readily. These rigid cells are
phagocytosed more readily than normal cells, producing
the characteristic anaemia.
1. Primary: the amino acid sequence of the protein linked

by peptide bonds.
2. Secondary: regular structures determined by hydrogen
bonding between the atoms of the backbone.
3. Tertiary: precise folding patterns of the polypeptide
chain stabilised by a wide range of bonds between the
amino acid side chains.
4. Quaternary: in multimeric proteins, the number and
arrangement of the subunits.
Primary structure
The primary structure of a protein is the sequence of amino
acids that makes up the protein chain. A chain with a large
number of hydrophobic residues will have completely differ
ent properties from one with many polar, charged side chains.
Likewise, the characteristics of a particular region of a protein
will depend on the average properties of the amino acids in
that region. Proline residues produce a very rigid molecule
with kinks in the protein chain, and glycine residues have
only a single hydrogen as their side chain and so take up
only a small space. Glycine and proline residues are, there
fore, often found in the regions of a protein where it loops
back on itself.
Secondary structure
The primary structure of a protein determines the overall
folding pattern of the polypeptide chain. However, there are
a number of regular patterns of folding which occur, stabil
ised by hydrogen bonding between the atoms of the back
bone of the peptide chain. These secondary structures are
not dependent on the nature of the side chains as the hydro
gen bonds form between the carbonyl group (C=O) of one
peptide bond and the hydrogen atom of another.
There are two common types of secondary structure
(Fig. 2.5), one forms a helical structure known as an a-helix
and another forms ribbons which can interact with one
another to form extensive b-sheets. A third important struc
tural motif is the U-turn, which allows protein chains to
rapidly change direction.
The a-helix
An -helix is formed when the polypeptide chain is twisted

into a rod, with the carbonyl group of each peptide bond
hydrogen bonded to the hydrogen of the amide, which is four
residues further down the chain (Fig. 2.5A). This coiling is
ch02.indd 23
Blood films: (A) normal; (B) with sickle cells.
always right-handed (clockwise) and has a pitch of 3.6 resi

dues per turn. Each amino acid adds 0.15nm to the length
of the helix. The structure is rigid and all the side chains face
outwards. This means that the overall hydrophobicity of
these -helical regions of the protein are determined by the
side chains and not by the backbone. Proline residues are
rarely found in -helices because their rigid structure would
not allow them to twist at the appropriate angle.
b-Sheets
-sheets occur where hydrogen bonds form between peptide

chains that lie alongside each other (Fig. 2.5B). Because the
four bonds around the -carbon atom are arranged in tetra
hedral fashion, the chains cannot lie completely flat and the
-sheet has a pleated appearance with the side chains pro
truding above and below the plane of the sheet. Some pro
teins are formed from multiple layers of -sheets stacked on
2/26/2009 6:48:29 PM
top of each other. An example of this is silk: its flexibility

results from -sheets sliding over one another, while its
strength is due to the axis of the protein backbone lying
parallel to the silk fibres.
Information box 2.2 a-Helical structures can

cross cell membranes
The -helix is commonly found in proteins that cross the

cell membrane. These proteins have regions in which there
are sequences of hydrophobic amino acid side chains that
are about 25 residues long. An -helix formed in these
regions gives a rod with a hydrophobic exterior about
3.75nm long, i.e. the length required to span the
hydrophobic interior of the cell membrane.
Some of the helices have hydrophobic residues on one
side of the helix and charged, hydrophilic residues on the
other side. These are called amphipathic helices and they
are found in proteins which form pores or channels in the
cell membrane.
A -Helix
H
U-turns
A third type of secondary structure formed due to hydrogen
bonding is the U-turn. A U-turn consists of three to four
residues, commonly glycine and proline, which form a short
loop often found on the surface of proteins. The absence of
a large side chain in glycine and the kink produced by proline
allow the polypeptide chain to fold tightly back in the protein,
facilitating a compact tertiary structure.
H
C
O
N
R
O
H
O
C
H
C
C
O
H
N
3.6 residues/turn
H
R
N
O
C
O
R
-Sheet
C
C
C N
C
R
H
C
C
C
C N
H
R
C
C
C N
H
R
C
C
C N
H
R
O
C N
C N
C
C N
C
C
C
C N
H
R
Fig. 2.5 Secondary structure of proteins: (A) -helix and (B) -sheet.
ch02.indd 24
2/26/2009 6:48:30 PM
Tertiary structure
Quaternary structure
The next level of three-dimensional organisation of the

polypeptide chain, the tertiary structure, results from a variety
of interactions between the amino acid side chains. These
interactions may include all the types of bonding already
described. Tertiary structure specifies exactly how the entire
protein is folded.
The type of interactions include:
Although some proteins consist of a single chain, there are

many multimeric proteins. These consist of multiple pro
teins held together by non-covalent bonds. These proteins
have an additional level of quaternary structure which speci
fies how many of each type of subunit is included (stoichi
ometry) and how they are arranged.
An example of this is the most common type of antibody
molecule, immunoglobulin G (IgG) (see Ch. 6). This type of
immunoglobulin is composed of four protein chains, called
heavy and light chains. The Y-shaped structure consists of
two heavy and two light chains, joined by disulphide bonds
and hydrogen bonds. Breaking the disulphide bonds between
the chains destroys the quaternary structure, and separately
the proteins chains have no biological activity.
Disulphide bonds, between cysteine residues on the

polypeptide chains
Hydrogen bonding between side chains that have
partial positive (+) and negative () charges, e.g.
hydrogen bonding between serine and asparagine side
chains
Ionic bonds between oppositely charged side chains,
e.g. between lysine and glutamate, sometimes called
salt bridges
Hydrophobic interactions in regions of the protein
where there are large numbers of non-polar, aliphatic
and aromatic side chains, which are excluded from the
aqueous surroundings.

Denaturation
The tertiary structure of many proteins can be disrupted by
heating, treatment with high concentrations of substances
such as urea, or reducing agents which can break disulphide
bonds. This is called denaturation and it leads to a loss of
both tertiary and secondary structure, abolishing the pro
teins biological activity (which itself is often used as an
assay of the degree of denaturation). The fact that some pro
teins can recover their activity when the denaturing agent is
removed shows that the tertiary structure of these proteins
is entirely determined by the primary structure. Examples of
such proteins are lysozyme (an enzyme found in tears and
saliva) and RNase (an enzyme which breaks down RNA).
Molecular chaperones
Not all denatured proteins can recover their structure and
activity. The recent discovery of proteins called molecular
chaperones or chaperonins, which aid the folding of the
protein chain, has shown that at least some proteins require
other factors to achieve the correct tertiary structure. These
chaperonins may also be involved in increasing the rate
at which naturally folding proteins can fold. Although many
proteins are able to fold correctly in vitro this is a slow
process and in vivo the chaperones make this process much
more efficient.
Information box 2.3 Hair is made up of many

a-helices
Hair is made up largely of a protein called keratin. Single

hairs are made up of hundreds of microfibrils embedded
in a protein matrix. Each microfibril is formed from a
number of -helices that are wound around each other
in a superhelix. Hairs can be stretched by elongating the
-helices, which breaks the hydrogen bonds in the helices.
When the hair is released it reverts to its previous length
because the covalent disulphide bonds between cysteine
residues in the -helix and the protein matrix remain intact.
Hair can be permanently curled or straightened by applying
chemicals which first break these disulphide bonds and
then re-form them in the new conformation.
ch02.indd 25
Protein modification
Almost all proteins are modified in some way after they have
been synthesised. Some of these modifications are perman
ent while others are reversible. Some involve chemical modi
fication of either the terminal carboxyl or amino groups
and/or modification of the side chains. Other changes
involve removal of regions of the protein after it has been
assembled.
Acetylation
The most common modification involves the addition of an
acetyl (CH3CO) group to the N-terminal. The effect of this
covalent change is to reduce the rate at which these proteins
are degraded, thus increasing their lifespan.
Glycosylation
Another common modification is the addition of carbohy
drate molecules to the surface of the protein, a process
called glycosylation. These glycoproteins and proteoglycans have many diverse roles including cell-to-cell recogni
tion and as components of the extracellular matrix.
Glycoproteins
Only serine, threonine or asparagine side chains can be
directly glycosylated. Serine and threonine are linked to
carbohydrates via their hydroxyl groups (O-glycosylation)
and asparagine via the amide group (N-glycosylation) of
the asparagine side chain. Another amino acid, lysine, can
have carbohydrates added to it after it has been modified
to hydroxylysine.
The carbohydrates found on glycoproteins are often
complex, branched molecules. Many of these glycoproteins
are components of the plasma membrane and have their
carbohydrate residues facing outwards and are thought to
be involved in cell-to-cell recognition processes.
Proteoglycans
The other types of protein which have carbohydrates linked
to them are called proteoglycans. They are protein chains
where the core protein is linked to large, linear carbohydrates
with a repeating unit.
Lipid modifications of proteins

Some proteins have fatty acids attached, which, through
their hydrophobic interactions can then be inserted into
the lipid bilayer of the cell membrane. These proteins are
thus concentrated at the inner face of the cell membrane
2/26/2009 6:48:30 PM

where they can interact more easily with other membrane
proteins.
Clinical box 2.1 Insulin secretion can be assessed

by measuring C-peptide
Phosphorylation
In patients with diabetes who need to inject insulin, the

presence of the injected insulin makes it difficult to measure
endogenous insulin release, because a direct insulin assay will
not be able to differentiate between the two sources of insulin.
However, for every endogenous insulin molecule released into
the blood, a C-peptide is also released and measuring the latter
gives an estimate of endogenous insulin production. Certain
conditions, however, such as a pancreatic -cell tumour, can
release large amounts of pro-insulin that will cause large errors
in insulin measurement.
The activity of many proteins can be reversibly modified by

phosphorylation. Enzymes called kinases add phosphate
groups to the hydroxyl groups of serine, threonine and tyro
sine residues found on many proteins. This change can have
the effect of increasing or decreasing the activity of the
protein. Other enzymes, phosphatases, can remove these
phosphate groups. These changes in structure can act as a
molecular switch that regulates the proteins activity.
Protein cleavage
Many proteins are formed from a precursor, which is then
modified to make the mature protein. An example of this is
the hormone, insulin, which is originally formed from a single
peptide chain with 84 amino acids (pro-insulin). After the
protein has been folded and disulphide bonds formed
between two parts of the chain, part of the central part of
the chain, called the C-peptide, is removed, leaving the two
ends of the polypeptide, called the A and B chains, con
nected by the disulphide linkages. If the protein is denatured,
for example by using mercaptoethanol which disrupts disul
phide bonds, the A and B chains are separated. When
the mercaptoethanol is removed the two chains do not reassociate, presumably because the most stable state for the
separate chains is different from that of the original single,
pro-insulin chain.
Enzymes
Almost all the biochemical reactions that take place in the
body require the presence of protein catalysts called
enzymes. Possibly as many as half of all proteins are
enzymes, each with their own unique structure. They are
globular proteins with irregular structures. They may contain
short stretches of -helices and -sheets, but their overall
forms vary enormously. Some enzymes fundamental to
general metabolism are found in all cells, whereas others
may only be found in specialised tissues. Enzymes act in the
same way as inorganic catalysts in that they lower the activ
ation energy of the reaction so that it can proceed under
physiological conditions, i.e. at temperatures and pressures
found in the human body. They can speed up the rate of
reaction between 106- and 1012-fold. Enzyme activity may be
increased in some diseases and reduced, or absent, in
others. Enzyme analysis can be an aid to the diagnosis of
many conditions.
The general mechanism of enzymic action is as follows:
1. The reactants called substrates bind to the enzyme to
form an enzymesubstrate complex.
2. The binding of the substrate causes changes in shape
to occur within the enzymesubstrate complex. These
internal rearrangements produce changes to bonds that
favour the formation of the product.
3. The product is released from the enzyme, which reverts
to its former shape. It can then go on to bind another
substrate molecule.
Specificity
The substrates bind to the enzyme at the active site by a
combination of ionic and hydrophobic interactions. The
ch02.indd 26
Proinsulin
S
S
S
S
S
S
Insulin
C-peptide
active site of each enzyme has a specific size and structure

that allows the substrate to fit into it exactly, a characteristic
that is often called a lock and key mechanism.
Some enzymes are very specific in their substrates, only
catalysing the reaction of a single substrate, whereas others
may accept a range of related substrates. Protease enzymes
break down proteins by breaking peptide bonds, and differ
ent proteases act on different proteins. For example, trypsin
is a proteolytic enzyme that will only hydrolyse those peptide
bonds to which arginine or lysine contribute the carboxyl
group. Pepsin, another gastric protease, is less specific in
its substrates and will hydrolyse a wider range of peptide
bonds.
Enzymes are very specific in the reaction they
catalyse. The same substrate can be converted to a number
of different products by the action of different reactionspecific enzymes. For example, pyruvate can be converted
to lactate, alanine, oxaloacetate or acetyl CoA by different
enzymes.
Enzyme nomenclature
Enzymes have specific names, depending on the reaction
catalysed, the substrates and the products. They are classi
fied into six classes, depending on the type of reaction cata
lysed (Table 2.7). However, many enzymes have a less formal
common name, which, for general use, describes their
action and often their substrate.
2/26/2009 6:48:30 PM
Table 2.7 Classes of enzymes
Class
Type
Action
Examples of common names
Oxidoreductases
Oxidationreduction reactions, often with coenzymes,

such as NAD+
Dehydrogenase, oxidase, peroxidase, reductase
II
Transferases
Transfer of amino, carboxyl, acyl, carbonyl, methyl,

phosphate groups from one molecule to another
Transaminase, transcarboxylase
III
Hydrolases
Cleave bonds between carbon and another atom by

inserting water
Esterase, peptidase, amylase, phosphatase,

pepsin, trypsin
IV
Lyases
Break carboncarbon, carbonsulphur and carbon

nitrogen (but not peptide) bonds
Decarboxylase, aldolase
Isomerases
Racemisation of optical or geometric isomers
Epimerase, mutase
VI
Ligases
Formation of bonds between carbon and oxygen,

sulphur, nitrogen, etc, often hydrolysing ATP (adenosine
triphosphate) to provide the required energy
Synthetase, carboxylase
Factors affecting enzyme activity
Substrate concentration
Enzyme activity (the rate of the enzyme-catalysed reaction)

is affected by a number of factors:
The other major factor which determines the rate of reaction

of an enzyme is the concentration of the substrate which is
written as [S], where the square brackets represent the con
centration. As the substrate concentration [S] is increased,
the rate of the reaction increases: at first the increase is linear,
then at higher [S] the rate of increase slows until the rate
reaches a plateau, where any further increase in [S] does not
produce any increase in the reaction rate.
This occurs because at low [S] the rate of reaction is
limited by the limited number of substrate molecules which
are bound to the active site. The initial increase in the rate
is due to the increase in the binding of the substrate to the
enzyme as the amount of substrate increases. As [S]
increases further, more and more of the substrate binding
sites are occupied until, at high [S] the enzyme is saturated
with substrate, i.e. all the binding sites are occupied by
substrate. At this point, even though there are more substrate
molecules available, there are no binding sites available. The
rate of the reaction is now limited by the rate at which the
enzyme can catalyse the reaction and the rate at which the
products leave the active site, making it available to more
substrate. The maximum rate of the reaction is called the
Vmax. Vmax is dependent on the quantity of enzyme present
and the conditions such as temperature and pH, which affect
the rate of catalysis.
Temperature
pH
Substrate concentration.

Temperature
Although normal body temperature is 37C, a large number
of enzymes increase their activity as the temperature is
increased. This is because of the increased kinetic energy of
the molecules and the increased likelihood the electrons will
be excited. If the temperature falls the rate of reaction falls
and sometimes the reaction will cease entirely. However, if
the temperature returns to normal, the enzyme activity is
regained.
If the temperature is increased above a certain level, the
protein melting point, the thermal excitation will be great
enough to break essential weak bonding forces and the
enzyme will become permanently disabled. At very high tem
peratures proteins become insoluble, as shown by the pre
cipitation of egg white proteins during cooking.
Even though they catalyse the same reaction, some
enzymes isolated from different tissues have different tem
perature resistance. For example, the differing denaturation
temperatures of lactate dehydrogenase (LDH) have been
used to distinguish heart and liver disease in assays where
LDH of cardiac origin appears more heat stabile than that
from the liver. The difference is due to LDH being present in
different forms.
Optimal pH
All enzymes have an optimal pH at which they function at
their fastest. Below or above this the enzymes activity is
reduced. For many enzymes the optimal pH is close to
pH 7. However, there are some enzymes which have their
optimal pH at very different acidities, reflecting the environ
ment in which they work. For example, the enzyme pepsin,
which digests proteins in the stomach has an optimal pH of
about 2 which is the pH of the gastric juice in the stomach
(see Ch. 15). Control of pH can also be a way of regulating
enzyme activity. Enzymes which are inactive under normal
conditions can become active when specific conditions
change the pH.
ch02.indd 27
Enzyme kinetics
If the initial rate of the reaction (v) at different [S] is plotted,
the resulting curve is a rectangular hyperbola, described by
the MichaelisMenten equation. This is derived from the rate
equation:
E + S ES E + P
where E is the enzyme, S is the substrate and P is the
product.
n = ( Vmax [ S ] ) ( Km + [ S ])
Vmax, the maximum reaction rate, can be estimated from
the plateau of the curve in Figure 2.6 where the rate of reac
tion is plotted against concentration. The other constant of
the MichaelisMenten equation is Km. This is defined as the
substrate concentration at which the rate is half Vmax. The
constant Km is informative, as it describes the ease with
2/26/2009 6:48:31 PM

which the substrate binds to the enzyme, or its affinity. Km
is inversely proportional to the affinity, with a low value of Km
indicating a high affinity. In other words, only a low concen
tration of substrate is required to saturate the enzyme and
vice versa. The two constants, Vmax and Km, are characteristic
for the enzymesubstrate combination under the conditions
under which they are measured.
Curve fitting and linear transformations

Without using statistical curve-fitting programs it is very dif
ficult to get accurate estimates of Vmax and Km from the
MichaelisMenten curve, especially as it is often difficult to
see where the plateau stops rising, which occurs at very high
levels of substrate. This leads to inaccurate estimates of Vmax
and because estimates of Km use this inaccurate value, they
themselves are inaccurate. This can be solved by using one
of two possible rearrangements of the MichaelisMenten
equation that produce straight lines when plotted, where the
kinetic constants can be easily identified from the slopes and
the intercepts. These plots are called LineweaverBurk and
EadieHofstee plots (Fig. 2.7 and Table 2.8). Both of these
linear transformations have some drawbacks because the
use of reciprocals tends to underestimate errors in the data
at different parts of the range. However, in the absence of
curve-fitting programs, they are much better than the stand
ard plot for estimating the kinetic constants.
The same mathematical models can be used to
describe:
Membrane transport (where Jmax is the maximum rate
of transport and Km the affinity of the transport system
for the substrate Carrier mediated transport)
Receptor binding (where the constants are called Bmax
and Kd, see Ch. 3).

tiple substrates can have different shaped curves, depending

on the mechanism by which the substrates bind. In particular,
reactions arising from cooperative interactions between sub
strate molecules, such as the binding of oxygen to
haemoglobin, have sigmoidal curves,
Enzyme inhibition
Many enzymes involved in metabolic pathways are inhibited
by the end-products or intermediates of those pathways.
This is an important mechanism for controlling the produc
tion of metabolites in appropriate amounts. Many drugs used
therapeutically act by inhibiting enzymes in order to reduce
the amount of a particular reaction product. Inhibition can be
reversible or irreversible.
Reversible inhibition
There are two types of reversible inhibition:

Competitive inhibition
Non-competitive inhibition.
They can be distinguished easily by looking at the effect

of the inhibitor on the kinetic parameters of the reaction.
Competitive inhibition
Competitive inhibition occurs because the inhibitor binds
reversibly to the same site on the enzyme as the substrate.
A
1
They can also be used to model drug interactions with a

variety of targets.
The MichaelisMenten equation applies only to a simple
reaction with a single substrate. Reactions that involve mul
1
1
Vmax
1
Km
[S]
Rate (V)
Vmax
(a and c)
a
Vmax
( b)
50% Vmax
(a)
0.5 Vmax
(b)
Vmax
Slope = Km
Km
Km
(a and b) (c)
Substrate concentration
[S]
Fig. 2.6 MichaelisMenten curves. Curves a and c have

the same Vmax, whereas curves a and b have the same Km.
[S]
Fig. 2.7 (A) LineweaverBurk plot: x-intercept = -1/Km,

y-intercept = 1/Vmax. (B) EadieHofstee plot: y-intercept =
Vmax, slope = -Km.
Table 2.8 Kinetic constants from different plots
ch02.indd 28
Type of plot
X-axis
Y-axis
Vmax
Km
MichaelisMenten
[S]
Plateau of curve
[S] at
LineweaverBurk
1/[S]
1/v
1/y-intercept
1/x-intercept
EadieHofstee
v/[S]
Y-intercept
slope
Vmax
2/26/2009 6:48:32 PM
Clinical box 2.2 Competitive inhibition of alcohol
dehydrogenase can be used to
treat poisoning by methanol and
ethylene glycol
Methanol is poisonous to humans and can cause blindness. It
can be formed accidentally in the production of homemade
spirits, or can be ingested intentionally as meths by alcoholics.
The metabolism of both methanol and ethanol (and the
antifreeze, ethylene glycol) involves the enzyme, alcohol
dehydrogenase. Part of the treatment of poisoning by both
methanol and ethylene glycol is the administration of ethanol in
order to block their breakdown, as it is the breakdown products
of methanol and ethylene glycol that are toxic. Ethanol has a
higher affinity (lower Km) for the active site of alcohol
dehydrogenase than either methanol or ethylene glycol and so
blocks their binding to the enzyme, and therefore their
subsequent breakdown. They can then be removed from the
blood by dialysis.
At low concentrations of substrate the enzyme activity is

reduced, but as [S] rises the inhibitor is displaced and at very
high [S] the enzyme activity returns to normal. This can be
seen by an increase in the Km of the enzyme; more of the
original substrate is now required to saturate the enzyme
than before, with Vmax unchanged. Competitive inhibition
often occurs between alternative substrates for the same
enzyme. The substrate with the lowest Km will be preferen
tially metabolised.
Non-competitive inhibition
Non-competitive inhibition occurs when the inhibitor binds
to a site on the enzyme other than the active site and by
doing so reduces the rate at which products are formed. The
binding of the substrate is unaltered so the Km remains the
same. However, at all [S] the rate of reaction is reduced, so
Vmax is reduced. Several chemotherapeutic drugs act in this
way.
Cooperativity between multiple subunits

In multimeric proteins cooperativity can occur between
multiple subunits. This happens when the binding of a mol
ecule, which can be a substrate or a regulator, to one subunit
induces a change in the quaternary structure of the protein,
which changes the affinity of the substrate-binding sites. This
means that small changes in the concentration of the mol
ecule cause a large change in the rate of catalysis.
For example, the oxygen-carrying protein, haemoglobin,
can bind four oxygen molecules. The binding of a single
oxygen molecule to one of the substrate-binding sites of
deoxyhaemoglobin (the deoxygenated form of haemoglobin)
causes a conformational change in the other subunits that
increases their affinity for oxygen so that the second oxygen
molecule binds more easily. This, in turn, increases the affin
ity of the other binding sites and so on, until all four sites are
filled. The reverse also occurs in that if one oxygen molecule
is unbound then the other molecules are lost more easily.
This cooperativity enables haemoglobin to become fully oxy
genated in the lungs and to lose this oxygen readily in the
tissues (see Ch. 13).
Isoenzymes
Many enzymes exist in different tissues in slightly different
forms, called isoenzymes. They may differ in their primary
structure or their subunit composition. Isoenzymes catalyse
the same reaction but may have different properties. For
example, they may have different affinities for their sub
strates. They may also show different patterns of inhibition
that help to regulate their function in different tissues.
Irreversible inhibition
Coenzymes
Irreversible inhibition occurs when the inhibitor is so tightly

bound to the enzyme that it cannot be removed and the only
way that the effect can be reversed is by the removal of the
enzyme and its replacement by newly synthesised enzyme.
For example, penicillin covalently inactivates a key enzyme
in bacterial cell wall synthesis.
Many enzymes require the presence of an additional compo

nent in order to function. These are called coenzymes,
Regulation of enzyme activity

Enzyme activity is regulated in several ways:
Feedback inhibition by the product of the pathway.
Many enzymes are inhibited by the products of their
metabolic pathways. The product of the pathway can
bind to a site on the enzyme and reduce the rate of
reaction.
Allosteric regulation: this is regulation by other small
molecules which can either activate or inhibit the
enzyme. The binding of the allosteric regulator to a site
other than the active site causes a change in the tertiary
structure of the protein, which alters the enzyme activity.
Gene expression: the amount of an enzyme is altered
by increased production of the enzyme in response to a
metabolic signal.

ch02.indd 29
Phosphorylation: as described earlier, the activity of

many proteins, including enzymes, can be reversibly
altered by the addition (with kinases) or removal (with
phosphatases) of phosphate groups.
Proteolysis: enzymes can be irreversibly activated or
inactivated by being broken down by other, proteolytic,
enzymes. The lifespan of enzymes can be changed by
altering their rate of proteolytic degradation in the cell.

Clinical box 2.3 Lactate dehydrogenase

isoenzymes can be used to
diagnose heart disease
Five different types of LDH are found in different human tissues.
Each type consists of a combination of four subunits of two
possible types called A and B. Type 1 consists of four B
subunits, type 5 of four A subunits with types 24 being
combinations of A and B subunits. The different subunits
determine how the enzyme is inhibited by its substrate, pyruvic
acid, and because of this, the degree to which the particular
tissue can undergo anaerobic respiration (i.e. without oxygen).
Heart muscle contains virtually only type 1, whereas skeletal
muscle and liver have mainly type 5. When someone has a
myocardial infarction (MI) and their heart muscle is starved of
oxygen, then muscle cells in the heart die and release their
contents into the blood. Analysis of the LDH isoenzymes in
the blood can confirm an MI as there would be more LDH-1
in the blood than normal. In contrast, in someone with liver
disease, you would tend to see increased levels of LDH-5 in
the plasma.
2/26/2009 6:48:32 PM

or cofactors. They may consist of metal ions such as
copper or iron, or they may be more complex organic mol
ecules. Some enzyme cofactors are derived from vitamins.
Vitamins are organic molecules that are required in small
amounts in the diet and without which deficiency diseases,
such as scurvy (vitamin C deficiency) or beriberi (vitamin B1
deficiency), can occur. Coenzymes are often involved in
oxidationreduction reactions, where they act as either elec
tron donors or acceptors. For example, the coenzymes,
flavine adenine dinucleotide (FAD) and nicotinamide adenine
dinucleotide (NAD), which are derived from vitamin B2
(riboflavin) and niacin, respectively, have a central role on
the transduction of energy in the electron transport chain
of mitochondria.
Structural proteins
Many of the proteins that form the structural elements of the
body are fibrous proteins made up of long repeating units,
arranged in bundles, which give strength and stability. These
make up the muscles, bones, skin and connective tissues.
They often have a simple repeated secondary structure of
-helices and -sheets, but because they are often mul
timeric they may also have a quaternary structure.
The three major groups of structural proteins are:
Collagens
Muscle proteins
Cytoskeletal proteins.
Information box 2.4 Mutations of type I collagen

cause brittle bone disease
(osteogenesis imperfecta)
There are many different forms of brittle bone disease, all

of which are characterised by weakness in all tissues
containing type I collagen. In each form there is a genetic
mutation that replaces the glycine in collagen with other
amino acids. These replacement amino acids are more
bulky and prevent the formation of stable triple helices and
collagen fibrils.
Table 2.9 Cytoskeletal proteins

Filament types
Main protein
constituent
Major role
Microfilaments
Actin
Cell movement
Form core of microvilli
Intermediate
filaments (IFs)
Keratins*
Mechanical strength
Microtubules
Tubulins
Chromosome separation
Intracellular transport
*The proteins making up IFs vary between cell types. They are made
up of keratins in many cells, but there are six different classes of IFs.
For example, nerve cells have IFs made from neurofilament proteins.
Collagens
The most abundant types of fibrous protein, which make up
about 25% of the total protein of the body, are the collagens.
They form the major component of the extracellular matrix
(ECM) which is the material that surrounds the cells. There
are a large number of different types of collagen (at least 20
have been described to date); they vary according to their
protein chains but they all have a similar quaternary
structure.
Collagen is made up of three polypeptide chains each of
which can be up to 3000 amino acids in length. Each chain
forms a left-handed helix that is tighter than the standard
-helix, with just three amino acids per turn. This tight helix
can be achieved because every third amino acid is a glycine,
which, with its single hydrogen, can fit into the core of the
molecule. The other amino acids are very often proline and
its derivative, hydroxyproline. The three helices are then
wound round each other into a right-handed triple superhelix,
which is stabilised by hydrogen bonding. The commonest
form of collagen (type I) can be found in a variety of tissues
and forms fibrils, with its molecules packed side by side that
confer great strength. Other types of collagen, such as type
IV, form sheets in which the molecules produce large flexible
networks. The basement membrane underlying epithelial
cells is partly made up of type IV collagen.
Muscle proteins
The main proteins that make up the contractile elements of
muscle cells are myosin and actin. They form the thick and
thin filaments of skeletal muscle, which interact to produce
muscle movement (see Ch. 9, Fig. 9.16). The two types of
filaments are formed in very different ways. Myosin is made
up of six polypeptide chains, two heavy chains and four light
ch02.indd 30
chains. The two heavy chains form -helices, which then coil
around each other. At their N-terminals they form two globu
lar head regions, with the four light chains. These globular
heads contain binding sites which interact with the actin and
adenosine triphosphate (ATP) to power muscle movement.
The strands of myosin aggregate into thick filaments made
up of 300400 myosin molecules.
The other filaments are composed of actin. Actin fila
ments are made by the polymerisation of molecules of globu
lar actin (G-actin) into long fibrous strands of F-actin. These
strands can be continuously extended and shortened by the
addition and removal of G-actin.
Cytoskeletal proteins
All cells contain cytoskeletal proteins that make up three
different types of filament, each made up of fibrous proteins
(Table 2.9). Each has a different role.
Other structural proteins

Elastin is a flexible protein, associated with collagen in
tissues which need to be elastic. It is found mostly in liga
ments and blood vessel walls, and is also found in skin,
tendons and connective tissues in much smaller amounts.
Many globular elastin molecules are cross-linked covalently
between lysine residues. When the tissue is stretched the
globular proteins can extend, but when the tension is released
they shorten back to their globular form. A mixture of colla
gen and elastin allows a certain degree of stretch, the degree
of which is controlled by the amount of collagen.
Two other groups of glycoproteins, important in forming
the basement membrane, the layer of material which lies
under sheets of cells, are laminin and fibronectin. These both
form part of the basal lamina, the part of the basement
membrane closest to the cell surface membrane. These mol
ecules can bind to proteins on the cell surface, as well as to
collagen and other ECM proteins.
2/26/2009 6:48:32 PM
Signalling proteins and receptors

Hormones and neurotransmitters
Some, but not all, of the signalling molecules which travel
from cell to cell, are proteins. These can be either hormones,
which are released into the bloodstream or neurotransmitters which are released from nerve endings.
An example of a protein hormone is insulin, which regu
lates blood glucose levels. Insulin is secreted from cells in
the pancreas into the bloodstream when blood glucose levels
are high and acts on other tissues to promote the uptake and
storage of the glucose. Hormonal responses are relatively
slow because hormones are often released at an area distant
from the site of action.
In contrast, when a rapid signal is needed, nervous
impulses can produce a rapid action through the release of
neurotransmitters (see Ch. 8) at the nerve terminals very
close to the site of action. In fact, many nerves secrete more
than one neurotransmitter. In response to low levels of stimu
lation, they secrete one of a number of small molecules, such
as acetylcholine or noradrenaline (norepinephrine). However,
in response to higher levels of stimulation, many nerves also
secrete a peptide neurotransmitter. An example of a peptide
neurotransmitter is the natural painkiller -endorphin. This
acts on opioid receptors, so called because they are also
activated by the opiate drugs, such as morphine.
Proteins are also involved in the signalling pathways
inside cells. For example, transcription factors are small
proteins which, when activated, bind to the genetic material,
DNA, to alter the type and amount of protein being produced
in the cell.
Receptors
Signalling molecules produce an effect on target cells by
interacting with proteins called receptors. The ability of a
cell to respond to any chemical signal (or ligand) depends
on the presence of receptors for that molecule. If the receptor
is not present then the cell cannot respond.
Cell surface receptors

For many ligands their receptors are embedded in the cell
membrane, with the binding site on the cell surface. Binding
of the ligand causes a conformational change in the receptor
that produces some effect inside the cell. This is called
signal transduction.
Intracellular receptors
Some signalling molecules, such as the sex hormones, tes
tosterone and oestrogen, can enter the cell and act on recep
tors present inside the cell. Some of these are present in the
cytoplasm, and others are located in the nucleus. The intra
cytoplasmic receptors have two binding sites, one for the
hormone and one for DNA. When the hormone is bound, the
receptor moves to the nucleus, binding with acidic proteins
of the DNA. The ligand-bound receptors, bound to the DNA,
produce changes in protein production, thus effecting cell
growth and metabolism.
Immune system proteins

Immunoglobulins
The body defends itself from attack by foreign bodies such
as bacteria and viruses with its immune system (see Ch. 6).
ch02.indd 31
Clinical box 2.4 The absence of testosterone

receptors produces babies that
look like girls who should have
been boys
Androgen insensitivity syndrome is a condition in which the
male fetus lacks the receptors for the male sex hormone,
testosterone. Genetically they are XY males, but a mutation
causes them to lack testosterone receptors. Despite producing
testosterone, these individuals cannot respond to it and do not
develop male genitalia because proteins necessary for the
appropriate cell growth and metabolism are not transcribed.
They do, however, produce normal amounts of other hormones,
which means that they develop external female features such as
breasts. Unfortunately, they do not develop internal female
organs, so cannot function as normal females.
One of the key elements of the immune system is the

production of vast numbers of different proteins called
immunoglobulins or antibodies which can bind to the in
vading organism or antigen, enabling the latter to be removed
and destroyed.
Antibodies have a common basic structure but include a
variable region which produces the binding site. These pro
teins are produced with an enormous variety of variable
regions to cope with the large number of possible exogenous
(i.e. of foreign origin) antigens. Problems can occur, however,
when the body produces antibodies that recognise its own,
endogenous, molecules as antigens. This failure to recognise
self results in autoimmune conditions, where the bodys own
cells are destroyed. A well-known example of this problem
is rheumatoid arthritis (RA).
Cytokines
Another group of proteins involved in both the immune
response and in the bodys response to inflammation are the
cytokines. More cytokines continue to be reported as new
research techniques are developed. Cytokines mainly act
locally, but some have been identified that have wide-ranging
effects on metabolism. One of the better-known groups of
cytokines are the interferons, which have actions against
viruses and cancer cells. One of them, interferon beta, is
currently being tested as a possible treatment for some
forms of multiple sclerosis.
Transport proteins
Many substances have to be transported around the body in
the extracellular fluids, and then subsequently into and out
of cells.
Transport in blood
Many proteins travel around the body in the fluid phase of
the blood: the plasma. Albumin is the major protein found
in blood plasma at a concentration of 3545g/L, making
up 50% of the total plasma protein. It has two major
functions:

To increase the osmotic pressure of the blood, thereby

ensuring the correct distribution of fluid between the
vascular and interstitial compartments (see Ch. 11)
To act as a non-specific transport protein for many
substances in the plasma.
2/26/2009 6:48:32 PM

A
Table 2.10 Some of the binding proteins in plasma

Protein
Ligand/s
Albumin
Metal cations, free fatty acids,

steroids, bilirubin, haem
Transferrin
Iron
Thyroid-binding globulin
Thyroxine (T4), triiodothyronine

(T3)
Cortisol-binding globulin
Cortisol
Sex-hormone-binding globulin
Androgens and oestrogens
O
C(1)
L-Glyceraldehyde
H Aldehyde
group
C(2)
OH
C(3)
OH
B
Albumin has an overall negative charge at physiological

pH, which means that it can easily bind metal cations such
as copper and iron. Many molecules, some of which are
extremely hydrophobic, can be transported in the aqueous
environment of the plasma, bound to albumin. This increases
the length of time they remain in the blood, due to reduced
degradation and elimination (see Ch. 2).
Other plasma proteins are specific binding proteins for
signalling molecules, such as sex steroids (Table 2.10).
D-Glyceraldehyde
H
C(1)
O Aldehyde
group
OH
C(2)
C(3)
OH
Dihydroxyacetone
H
H
OH
OH
Keto
group
Fig. 2.8 Triose sugars: (A) aldoses and (B) ketoses.
Transport into cells

Hydrophobic substances can move into cells by simple dif
fusion but many substances, including glucose, cannot move
into cells without the presence of specific proteins called
transporters. These transporters are proteins embedded in
the cell membranes that bind the transported molecule on
one side of the membrane and move it through to the other
side. Inside cells, substances can be moved into separate
compartments by the action of other specific transport
proteins.
CARBOHYDRATES
Carbohydrates form about 2% of body mass. They consist
of carbon, oxygen and hydrogen, with a ratio of hydrogen
to oxygen of about 2:1, hence the name hydrated carbon.
The most common organic compound on the planet is a
carbohydrate, cellulose, which forms the principal compo
nent of plant cell walls. Unfortunately, humans cannot digest
significant amounts of cellulose, although it is important to
digestion as a bulk element of food. Of more importance is
the carbohydrate, glucose, which is one of the central mol
ecules in energy metabolism and forms the backbone for the
synthesis of many other compounds. The simplest forms
of carbohydrates are called sugars. These are either mono
saccharides, which are single units or disaccharides,
formed from two monosaccharides. Larger carbohydrates,
called polysaccharides are polymers, formed from many
monosaccharides.
Sugars
Monosaccharides
The basic units of all carbohydrates are called monosaccharides, or simple sugars. These can be obtained directly
from the diet or derived from the breakdown in the gut of
more complex carbohydrates. They can also be manufac
tured from non-carbohydrate sources when carbohydrate
sources have been used up. Although food is the main
source of sugars there is no dietary requirement for carbo
hydrates as such.
ch02.indd 32
Triose sugars
Sugars have the general formula (CH2O)n where n is the
number of carbons in the sugar. The simplest of these are
the three-carbon trioses. Two types of triose sugars contain
either an aldehyde group, aldoses, or a ketone group, ketoses
(Fig. 2.8). In aldose sugars, the carbon atom of the terminal
aldehyde group is numbered carbon-1. In ketose sugars,
carbon-1 is the end carbon next to the carbonyl group. The
central carbon of glyceraldehyde, the simplest of the aldose
sugars, is chiral, and therefore it can exist as two stereo
isomers, d- and l-glyceraldehyde.
d
and
sugars
All sugars larger than trioses have two or more chiral groups,
and so exist as a number of stereoisomers. By convention
they are divided into d- and l-series, depending on the
arrangement of atoms around the asymmetrical carbon fur
thest from the carbon-1 end. The sugar is a d-sugar if
the arrangement around this carbon is the same as dglyceraldehyde, and an l-sugar if it is like l-glyceraldehyde.
Virtually all sugars found in biological systems are
d-isomers.
Different forms of glucose

d-Glucose, also known as dextrose, is found in all cells and
in blood plasma, where its concentration is strictly controlled.
Glucose is a six-carbon sugar, or hexose, and has the
formula, C6H12O6. It can exist in four different forms, all of
which are freely convertible in aqueous solution (Fig. 2.9).
The straight chain form (which exists as less than 1%) has
an aldehyde group at one end (where the carbon is always
called carbon-1). This reacts with the hydroxyl group on
carbon-5 to give a six-member ring form called a pyranose
ring. Depending on the arrangement of the H and OH
on carbon-1, the glucose molecule forms either - or
-glucose (also known as -d-glucopyranose and -dglucopyranose). These different forms are known as
anomers. While the cyclic form of d-glucose is usually a
pyranose ring, rarely it can also form a five-member furanose ring, d-glucofuranose.
2/26/2009 6:48:32 PM
A
Straight chain
OH
O
C(1)
C(2)
H
H
Aldehyde
B
H
C(3)
OH
C(4)
C(5)
CH2OH
H
(6)
OH
OH
-D-Glucose (-D-Glucopyranose)
H
C(4)
OH
C(5)
H
OH
C(3)
C(2)
OH
OH
OH
CH2OH
C(1)
OH
H
OH
OH
OH
H
OH
-D-Glucose (-D-Glucopyranose)
H
C
OH
-D-Galactose
CH2OH
O
H
OH
OH
CH2OH
OH
D-Glucofuranose
CH2OH
OH
C(1)
HCOH
C(4)
H
OH
OH
-D-Fructofuranose
H
O
(5)
OH
Fructose
(6)
OH
C(3)
C(2)
C(1)
OH
OH
H
OH
Fig. 2.9 Different forms of glucose. Each of these
molecules has an identical chemical composition. A
reaction between the hydroxyl group on C5 and the
aldehyde on C1 produces the pyranose ring structure (B,
C). If the hydroxyl group is on C4 then the furanose ring is
produced (D).
OH
OH
Due to the tetrahedral arrangement of bonds around the

carbon atoms, the pyranose and furanose rings are not
planar molecules. They form two different arrangements,
known as the chair and boat conformations. Both of
these conformations are important for the packing of the
molecules, although the chair conformation is the more
stable.
Common hexose sugars

Other common hexoses are mannose, galactose and fruc
tose (also known as fruit sugar) (Fig. 2.10). Mannose
and galactose are both aldoses and tend to form pyran
ose rings. They are only different from glucose in the
configuration of the other carbons, and are called epimers.
Mannose is identical to glucose, except for the arrange
ment around the carbon-2. Similarly, galactose only differs
from glucose by the arrangement around carbon-4.
Fructose is a ketose sugar and forms both furanose and
pyranose rings.
CH2OH
OH
CH2OH
H
CH2OH
OH
OH
OH
D-Ribose
CH2OH
H
OH
H
OH
ch02.indd 33
-D-Mannose
CH2OH
CH2OH
(6)
-D-Glucose
A
Hydroxyl
OH OH
OH
Deoxyribose
O
CH2OH
H
OH
OH
Fig. 2.10 Structures of some common sugars. In (B)

and (C) the part of the molecule which differs from glucose
is highlighted.
Pentose sugars
Two important pentose sugars are ribose and deoxyribose,
which are found as components of the nucleic acids, DNA
and RNA. These are both aldoses and form cyclic furanose
rings.
2/26/2009 6:48:33 PM

Table 2.11 Principal disaccharides
Disaccharide
Carbon-1 sugar
Other sugar
Linkage
Digestive enzyme
Sucrose
-Glucose
-Fructose
12
Sucrase-isomaltase
Lactose
-Galactose
-Glucose
14
Lactase
Maltose
-Glucose
-Glucose
14
Maltase
Isomaltose
-Glucose
-Glucose
16
Sucrase-isomaltase
Other simple sugars

Other carbohydrates can be derived from basic carbohy
drates by the inclusion of other elements. Glucosamine and
galactosamine are derived from glucose and galactose,
both having amino groups replacing the hydroxyl groups on
carbon-2. These two sugars are major components of the
proteoglycans, which make up the ECM.
Among other possible substitutions are:

Phosphates widespread in energy metabolism

Sulphates and carboxylates: commonly found in
proteoglycans.
Disaccharides
Monosaccharides can be linked together by glycosidic linkages to form disaccharides. These links are formed by a
dehydration reaction between the hydrogen on carbon-1
atom of one sugar and the hydroxyl group of another, with
the loss of a water molecule. This reaction is reversible.
These are either or linkages, depending on the orientation
around carbon-1 atom. This means that there are a large
number of different possible bonds that could occur between
two sugars, each of which gives a different product (Table
2.11). For example, glucose has five different OH groups
(on carbons 1, 2, 3, 4 and 6), so could potentially form dimers
in 10 different conformations. If two different sugars are
combined the second sugar could be either an -sugar or a
-sugar, increasing still further the possible combinations.
However, enzymes are able to distinguish between the
different conformations ensuring that the required linkage
is either made or broken in order to produce the required
product.
Dietary sugars
The compound known as ordinary sweet-tasting sugar is
actually a disaccharide, sucrose, which is made up of glucose and -fructose in an (12) linkage. Two other
important dietary disaccharides are maltose, which is a
dimer of two glucose molecules, and lactose (called milk
sugar, because milk is its only significant source), which is
made from galactose and glucose. Dietary disaccharides are
hydrolysed by specific enzymes found in the intestine. The
resulting monosaccharides are then absorbed by specific
transport mechanisms (see Ch. 15).
Polysaccharides
Polysaccharides are polymers of monosaccharides and
may contain thousands of units. They allow large amounts
of glucose to be stored in a highly concentrated form, without
the osmotic problems that would be associated with high
aqueous concentrations of separate glucose molecules.
Starches
These are a group of polysaccharides which are used as
sugar storage in plants. They consist of polymers of glucose
ch02.indd 34
Clinical box 2.5 In certain populations adults

cannot digest lactose
The enzyme, lactase, which breaks down lactose into galactose
and glucose, is usually present only in children and only
persists in adults of northern European descent. Lactase
deficiency is common in Mediterranean countries, parts of
Africa and Asia. In the absence of lactase, consuming milk
products containing lactose produces watery diarrhoea,
flatulence and abdominal pain. This is because the unabsorbed
lactose is fermented by bacteria in the small intestine,
producing gas and large numbers of metabolites that draw
water into the intestine by osmosis, producing diarrhoea. This
can be avoided to some extent by fermenting the milk, to
produce yoghurt, for example, because the lactose is converted
to lactic acid during fermentation.
linked in two different ways. The simplest starch, amylose,

consists of long linear chains of glucose molecules, linked
via 14 bonds. Amylopectin, which makes up about 80%
of the starch in foods, also has 14 linked glucose chains,
but additionally every thirtieth glucose molecule is also linked
via a 16 bond, producing a branched molecule. The
starches found in grains, such as wheat or rice and potatoes,
are the major source of dietary carbohydrates. They can be
broken down to glucose in the human gut by amylases
secreted by the pancreas and the salivary glands. These
enzymes break starches into glucose, maltose and isomal
tose (from the 16 linkages in amylopectin).
Non-starch polysaccharides
Cellulose is an unbranched glucose polymer with 14
linkages. Humans have no enzymes capable of hydrolysing
these linkages, although a small amount of cellulose is
broken down by bacteria in the colon. However, cellulose,
along with other non-metabolised polysaccharides, are the
major component of dietary fibre. They produce bulk in the
intestine which aids defecation.
Glycogen
Glycogen is the storage form of glucose in animals, including
humans. Like the starches, it is a large glucose polymer. It is
similar in structure to amylopectin, except that the branch
points occur every tenth glucose molecule. The high level of
branching of the molecule means that it has a large number
of free ends to allow glucose molecules to be added or
removed rapidly. Glycogen stores are not very large in
humans, with about 75g in the liver and about 250g in
skeletal muscle. Each gram can produce 16.7kJ (4kcal), so
glycogen stores can provide sufficient glucose for 1218
hours in the absence of other sources of glucose. Dieting
rapidly depletes glycogen, with an apparent rapid weight
loss. However, this loss is mainly due to the loss of water
molecules that are associated with glycogen.
2/26/2009 6:48:33 PM
Table 2.12 Major glycosaminoglycans (GAGs)
GAG
Main source
Hyaluronic acid
Joints
Chondroitin sulphate
Cartilage
Keratan sulphate
Cornea
Dermatan sulphate
Skin
Heparan sulphate
Basement membranes
Heparin
Mast cells
Functions of carbohydrates
Glucose, as mentioned above, is a major metabolic fuel, and
it is the only fuel used by the brain under normal circum
stances. Red blood cells are completely reliant on glucose
for energy because they lack the ability to metabolise fats.
Also, at the beginning of exercise, muscle preferentially uses
glucose rather than other fuels. Glucose is oxidised by cells
in a process called glycolysis, which liberates 16.7kJ (4kcal)
per gram of glucose. The intermediates of glycolysis can also
provide precursors for many amino acids, nucleotides and
lipids.
Other functions of carbohydrates include forming struc
tural elements surrounding cells and playing a part in the
mechanisms by which cells recognise each other.
Structural carbohydrates
The proteoglycans, which form the gel part of the ECM,
typically contain 95% carbohydrate. They are formed from
two components: a linear polypeptide chain, to which are
attached large, linear polysaccharide chains called glycosaminoglycans (GAGs) or mucopolysaccharides (Table
2.12). These GAGs consist of repeating disaccharide units.
Many of these are negatively charged and thus attract cations
and large amounts of water, drawn into the gel by osmosis.
This results in a gel-like structure around the collagen fibres
of the ECM, imparting a degree of flexibility to the tissue and
acting as a shock absorber. In contrast, the GAG, heparin,
although having a similar structure to the other GAGs, has a
very different function. It is released from mast cells and acts
as an anticoagulant (see Ch. 12).
Cell recognition by cell surface glycosylation

The carbohydrates found in glycoproteins are of two types.
One type, which consists of one or two simple mono- or
disaccharides, are linked to serine and threonine residues
by O-glycosylation. The linkage is between the hydroxyl
group of the amino acid side chain and the carbon-1 of
N-acetylgalactosamine.
The other way that sugars are linked to proteins is via Nglycosylation to asparagine. This always occurs at specific
amino acid sequences in the protein, either asparagine-Xserine or asparagine-X-threonine, where the linkage is
between the amide group of asparagine and the carbon-1
of N-acetylglucosamine. Monosaccharide units are then
added, in a specific pattern, to give highly branched carbo
hydrates. Many of these carbohydrates contain the sugar
N-acetylneuraminic acid (sialic acid). This sugar is negatively
charged, giving many cell surfaces a net negative charge.
Lipids in the cell membrane are also glycosylated.
These distinctive patterns of glycosylation are particularly
important in cell-to-cell recognition and how the immune
system recognises self. An example of this specificity can be
seen on red blood cells.
ch02.indd 35
Blood groups
The surface of the red cell is covered with patterns of car
bohydrates attached to protein and lipids, making the many
different antigens that determine different blood groups.
The most studied of these is the ABO blood group system.
The O antigen is a small polysaccharide, present on all red
cells of all blood groups. The A and B antigens are produced
by the addition of N-acetylgalactosamine or galactose sugar,
respectively, to this O antigen. These are added by specific
transferase enzymes; individuals with the AB blood group
have both enzymes and their erythrocytes have both A and
B antigens (see Ch. 12).
LIPIDS
In adults who are not overweight or underweight, lipids make
up about 20% of body mass, usually more in women. Unlike
proteins, carbohydrates and nucleic acids, lipids are not
large covalently bonded molecules. However, because of the
interactions that occur between lipid molecules, they form
large structures of repeating molecules. Lipids have a number
of crucial roles:

They form the membranes surrounding every cell and

the compartments within the cells.
They are also the major form of energy storage.
They have important roles in cell signalling.
Abnormalities in lipid transport are associated with the
development of atherosclerosis, where arteries are clogged
with fatty deposits, and excess storage of lipids results in
obesity. Diseases such as diabetes mellitus and pancreatitis
have associated lipid abnormalities, and there are also rare
inherited lipid storage diseases.
Triacylglycerols
The simplest type of lipids, the triacylglycerols, also called
triglycerides or neutral fats, are formed from two compo
nents, fatty acids and glycerol (Fig. 2.11). Triacylglycerols
have three ester bonds, formed by dehydration reactions
between the carboxylate group of three fatty acids and each
of the three hydroxyl groups of glycerol (Fig. 2.11D).
Fatty acids
Fatty acids are chains of carbon and hydrogen (Fig. 2.11A,
B), with a carboxylate (COOH) group at one end (the
-carbon) and a methyl group (CH3) at the other (the
-carbon):

If all the carbon atoms in the chain are single bonded

then the fatty acid is saturated.
If there is a single double bond it is monounsaturated.
If there is more than one it is polyunsaturated.
Most of the double bonds in biological molecules are in
the cis-configuration, which puts a rigid kink in the hydrocar
bon chains. Many of the fatty acids found in cells have 16,
18 or 20 carbon atoms and up to three double bonds.
Glycerol
Glycerol is a sugar alcohol, a modified triose sugar
formed by the reduction of the aldehyde group to hydroxyl
(Fig. 2.11C).
2/26/2009 6:48:33 PM

Saturated fatty acid
OH
C()
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CH2
()
CH2
CH2
CH3
CH2
Monounsaturated fatty acid (oleicacid, C18: 19)
B
OH
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CH
CH
CH2
CH2
CH2
CH2
CH2
CH2
CH3
(9)
Glycerol
H
OH
OH
OH
Triacylglycerol
H
O
H
C
O
CH2
C
O
CH2
CH2
Fig. 2.11 Components of triacylglycerols.
Information box 2.5 Fatty acids can be named

in several ways
Many of the different fatty acids have common names

which reflect their original source, but all fatty acids can be
described according to the number of carbon atoms in the
chain and the number and position of the double bond/s.
A shorthand notation that is widely used gives the
number of carbon atoms, the number of double bonds
and the position of the first double bond, counting from
the -carbon. For example, the saturated fatty acid
found in palm oil is called palmitic acid. The formula for
palmitic acid is CH3(CH2)14COOH, its chemical name is
n-hexadecanoic acid and the shorthand notation is C16:0,
indicating that it has 16 carbon atoms and no double
bonds.
Another fatty acid, arachidonic acid, is called cis5,8,11,14-eicosatetraenoic acid, which is shortened to
C20:4 -6. Another type of shorthand indicates the
positions of all of the double bonds: C20:4 All cis5,8,11,14 (see Fig. 2.14).
Dietary fats
Dietary fats, both solid and liquid (oils), are mainly com
posed of triacylglycerols. The fatty acids which make up the
triacylglycerol molecules in each type of fat determine the
physical properties of the fat. If the triacylglycerol has fatty
acid chains that are predominantly short, or contains unsatur
ated fatty acids, it will be liquid at room temperature. Exam
ples of these are the oils such as olive oil (which contains
oleic acid) and sunflower oil, which contain polyunsaturated
fatty acids. If there are more saturated fats and longer
fatty acid chains then the resulting fat will be solid at room
temperature, for example, butter and the fats found in
meat.
ch02.indd 36
Formed from glycerol and

3 molecules of palmitic acid
Adipose tissue
Triacylglycerols tend to aggregate with the formation of hydro
phobic bonds between them which exclude water. This
can be seen clearly when oils are added to water. They do
not mix and form droplets. Instead they form a thin layer
over the surface. In the body, triacylglycerols form the major
energy store. They are stored in white adipose tissue. This
is found mainly under the skin, and also surrounding the
abdominal organs and on the hips. Adipocytes, specialised
mesenchymal cells, each consist almost entirely of a single
droplet of triacylglycerols. The molecular aggregation of the
lipid molecules means that the lipids do not have an osmotic
effect; i.e. despite the presence of large numbers of mol
ecules, they do not act like individual particles. Obesity is
characterised by increased size and number of these cells.
Fats yield 37.7kJ (9kcal) per gram and the amount of fat
stored in the adipose tissue in a lean average human would
be enough to survive for more than two months without food.
There is no specialised storage site for proteins or amino
acids. In severe starvation the body will break down tissue
and plasma proteins in order to produce fatty acids.
Glycogen also has no specialised storage cell; it is simply
accumulated in the cytoplasm, particularly in liver and
muscle.
Membrane lipids
The lipids which form cell membranes are of three types:

Phospholipids
Sphingolipids
Cholesterol.

Two of these have broadly similar structures. These are

the glycerolipids, which are mainly phospholipids, and the
sphingolipids.
2/26/2009 6:48:34 PM
A
Phosphatidylserine
Phosphate
O
COO
NH3+
CH
CH2
Serine
CH2
CH
Hydrophilic
CH
O
Glycerol
R = Fatty acid hydrocarbon chains
Hydrophobic
Sphingomyelin
CH3
CH3
N+
Phosphocholine
CH2
CH2
O
P
O
CH3
CH2
Sphingosine
NH
Hydrophobic
CH
CH
(CH2)12
CH3
OH
C
Hydrophilic
R = Fatty acid hydrocarbon chains
Fig. 2.12 Two membrane lipids.
Types of membrane lipids
Phospholipids
CH2
Phospholipids (Fig. 2.12A) consist of glycerol, with fatty

acids attached to two adjacent OH groups via ester linkages
to give two acyl groups, and with a third group attached,
which is a phosphate group. The phosphate is itself linked
to one of a number of possible molecules, including serine,
ethanolamine (a derivative of serine), choline and inositol.
The resulting molecular structure looks somewhat like a
tuning fork, with hydrophobic prongs and a hydrophilic
handle.
Sphingolipids
Sphingolipids (Fig. 2.12B) have a similar structure to
phospholipids, but the hydrophobic portion is formed from
sphingosine. Sphingosine contains a long hydrocarbon
chain and as part of a sphingolipid it is linked to a fatty
acid, giving the sphingolipid two long hydrocarbon chains.
This in then linked to either a phosphate-containing group,
phosphocholine, or a group of sugar residues to produce
a glycolipid.
Cholesterol
A third component of cell membranes is cholesterol. This
has a structure completely different from the other mem
brane lipids (Fig. 2.13), as it is formed from a four-ringed
hydrocarbon called a steroid. Although cholesterol is almost
entirely hydrocarbon, it has a hydroxyl group attached to the
first ring, which can interact with water. The ring structure
is quite rigid and has important effects on the fluidity of
membranes.
Arrangement of membrane lipids

All the membrane lipids are amphipathic; that is they have
a hydrophilic head group and a hydrophobic tail. They aggre
ch02.indd 37
CH2
CH3
CH3
CH2
H
CH3
CH3
Hydrophobic
CH3
Hydrophilic
Steroid
OH
Fig. 2.13 Structure of cholesterol. The figure inside the

dotted blue lines is the basic steroid structure.
gate with their head groups facing the aqueous medium and
their tails excluding water. It is this arrangement which under
lies the formation of all cell membranes (see Fig. 2.28). Many,
although not all, of the head groups are negatively charged,
giving the surface of the aggregated lipids an overall negative
charge.
Different membranes vary in their relative composition.
For example, the myelin sheath around nerves has a very
high concentration of sphingolipids, whereas phospholipids
are more prevalent in the red cell membrane.
An alternative arrangement of amphipathic lipids in
aqueous solution leads to the formation of spherical micelles
(see Ch. 15), structures generated, for example, by bile salts
during the digestion of dietary fat.
2/26/2009 6:48:34 PM
Cholesterol and other steroids

Cholesterol is the major sterol (a steroid alcohol) in the
human body, having both structural and transport functions.
It can be synthesised in the liver or absorbed in the diet.
Studies have shown a relationship between raised plasma
cholesterol (in the form of low-density lipoprotein (LDL)cholesterol; see Ch. 16) and increased mortality from heart
disease. It is therefore thought to be beneficial to reduce
plasma cholesterol levels. However, reducing dietary choles
terol has less of an effect on plasma cholesterol levels than
reducing the intake of saturated fats. Diets that are rich in
polyunsaturated fats can also reduce serum cholesterol.
As well as forming an important part of membranes, chol
esterol is the precursor of many important molecules that are
formed by modifications of the basic steroid structure (see
Fig. 2.13). These are:

Bile salts
Steroid hormones
Vitamin D.
Bile salts
Bile salts act as detergents, helping to break up fats, so
aiding their digestion and absorption. They are produced in
the liver and are secreted from the gall bladder into the
intestine (see Ch. 15).
tradition of not exposing the skin, then dietary sources of

vitamin D are important (see Ch. 16).
Essential fatty acids

While most of the fatty acids used in the body are supplied
in the diet (Fig. 2.14), there is almost no requirement for any
fat in the diet. Saturated fatty acids can be synthesised from
the breakdown products of carbohydrates and proteins in a
process called lipogenesis (see Ch. 3). The requirement for
mono- and polyunsaturated fatty acids is met by enzymes
that convert saturated fatty acids to unsaturated fatty
acids.
However, some fats are required:

To aid the absorption in the gut of fat-soluble vitamins

As precursors to the eicosanoids.
These essential fatty acids are linoleic acid and linolenic acid, both of which were originally identified in linseed
oil. They cannot be manufactured in the body as none of the
enzymes which convert fatty acids can include double bonds
beyond carbon-10, and an important group of lipids, the
eicosanoids are all made from 20 carbon fatty acids with
between three and five double bonds. Arachidonic acid, the
precursor for a large number of different eicosanoids, is syn
thesised from linoleic acid (see below).
Eicosanoids
Steroid hormones
Steroid hormones are all derived from cholesterol, which is
converted to pregnenolone and then into the hormone impor
tant in maintaining pregnancy, progesterone (Table 2.13).
This can then be further modified to produce all of the other
steroids (see also Ch. 10). Although they are quite large
molecules, because they are lipid soluble, they can cross
plasma membranes and act on receptors found inside cells.
They are usually carried in the blood, associated with binding
proteins such as albumin (see Table 2.10).
These are a large group of locally acting hormones derived

from fatty acids (Fig. 2.14). They are extremely potent sub
10
6
13
11
12
4
14
Group
Example
Major actions
Glucocorticoids
Cortisol
Formation of glucose,
breakdown of proteins
and lipids
Mineralocorticoids
Aldosterone
Sodium and potassium

homeostasis
Progestogens
Progesterone
Pregnancy and ovarian

cycles
Androgens
Testosterone
Male sexual
development and
characteristics
Oestrogens
ch02.indd 38
Oestradiol
Female sexual
development and
characteristics
16
15
(1)
18
17
20
19
PGE2 (prostaglandin E2)

O
COOH
OH
Table 2.13 Important steroid hormones
COOH
Point of
cyclixation
Vitamin D
The precursor of vitamin D is synthesised in the skin from
cholesterol, where it is converted by the action of ultraviolet
light to cholecalciferol. Further processing of this molecule
produces active vitamin D which acts to influence the
absorption of calcium in the intestine, reabsorption in the
kidney and bone resorption and calcification. When skin
exposure to light levels are low, such as might occur in
countries outside the tropics in combination with a cultural
Arachidonic acid
OH
PGI2 (prostacyclin)
COO
O
OH
OH
Fig. 2.14 Arachidonic acid. All cis, 5, 8, 11, 14
eicosatetraenoicacid (or alternatively C20: 46 C-20 5,

8, 11, 14) and two of the many eicosanoids derived
from it (PGE2 and PGI2 (prostacyclin)).
2/26/2009 6:48:34 PM
Information box 2.6 Prostanoids facilitate normal

blood flow through vessels
and aspirin acts by inhibiting
the synthesis of
prostaglandin
Blood is protected from clotting in blood vessels, under

normal conditions, due to the release of a compound,
prostacyclin, from the vessel wall. Prostacyclin acts as a
vasodilator, thus encouraging flow, and inhibits the sticking
together of blood platelets, which form the basis of clot
formation. If the vessel wall is broached substances are
released that stimulate platelets to release another
prostaglandin, thromboxane A2. In contrast to prostacyclin,
thromboxane A2 acts as a vasoconstrictor, thus reducing
blood loss through the leaking vessel wall, and stimulates
platelets to aggregate, plugging the hole in the vessel.
The enzyme which acts on arachidonic acid in the first
step of the production of the prostanoids is cyclooxygenase (COX). Aspirin (acetylsalicylic acid) is one of a
group of drugs called non-steroidal anti-inflammatory drugs
(NSAIDs) which inhibit COX and reduce inflammation and
pain. Aspirin is also used to inhibit blood clotting by daily
consumption of a low-dose, since, in the absence of COX,
thromboxane A2 cannot be produced. Because of the
enzyme inhibition by NSAIDs, gastric bleeding is seen as a
common side effect. COX is seen in two isoforms, COX-1
and COX-2. While COX-1 is continuously produced, the
other isoform, COX-2, is only induced in response to
inflammatory mediators. Although current NSAIDs inhibit
both COX-1 and COX-2, it is thought that if specific COX-2
inhibitors were developed, they might be able to reduce
inflammation without the side effects of current NSAIDs.
stances and are all broken down quickly after being released,
so they act only over short distances. They have important
roles in responses to inflammation and in the control of physi
ological processes, showing a wide range of functions. For
example, some stimulate smooth muscle cell contraction,
producing vasoconstriction, while others act as vasodilators.
The different groups of eicosanoids are:

Prostaglandins
Thromboxanes (together these are called the
prostanoids)
Leukotrienes.

Arachidonic acid, or eicosatetraenoic acid, is one of

the fatty acids incorporated into membrane phospholipids. It
can be released from the membrane by the action of the
enzyme phospholipase A2 (PLA2) on phosphatidylcholine,
one of the most common membrane phospholipids. PLA2 is
stimulated by a range of different signals in different tissues,
including general cell damage.
Another fatty acid, eicosapentaenoic acid, is the pre
cursor to a group of prostaglandins with three double bonds.
One of the double bonds is at carbon-17, which is three
carbons from the -carbon, hence the name of this type of
fatty acid which is -3 fatty acids. It is thought to be bene
ficial to include this polyunsaturated fatty acid, which is
found in fish oils, in the diet.
NUCLEIC ACIDS
Deoxyribonucleic acid (DNA) and the ribonucleic acids
(RNAs) contain and transmit the genetic information in
all biological organisms. Nevertheless, they are not very
complex, in the sense that they are regularly arranged
ch02.indd 39
Base
Phosphate
Sugar
NH2
N
O
O
P O
CH2
O
Phosphate
N
H
OH
N
Adenosine
OH
Ribose
Fig. 2.15 Nucleotide structure: (A) components of a

nucleotide; (B) structure of ribo-adenosine monophosphate
(rAMP), usually referred to more simply as adenosine
monophosphate.
molecules made from a limited number of monomers. Nucleic

acids, like proteins, are unbranched chains of units that are
all linked with a similar type of bond. However, instead of the
20 possible different subunits which can make up proteins,
each of the nucleic acids has only four different subunits. In
this way they would seem to be much simpler molecules than
proteins, but the way in which the nucleic acid chains are
used to store and transmit the genetic information of every
cell indicates that the information contained within these
macromolecules is complex (see below).
In addition to their genetic role, some of the subunits of
nucleic acids also have important roles in energy transfer and
as cell signalling molecules.
Nucleotides
Nucleotides form the basic units from which the larger
nucleic acids are made. Nucleotides have three components
(Fig. 2.15):

A sugar
A phosphate
A base.

Nucleotide sugars and phosphates

Two different sugars are found in nucleotides, deoxyribose
and ribose. These pentose sugars are both aldoses with a
furanose ring (see Fig. 2.10E,F).
Attached to the hydroxyl on carbon-5 of these sugars by
an ester bond is one of three possible phosphate-containing
groups. These phosphate-containing groups can have either
one, two or three phosphates attached to each other via
phosphoanhydride (pyrophosphate) bonds between the
phosphates.
Nucleotide purines and pyrimidines

There are two types of bases found in nucleotides: the
purines and the pyrimidines (Fig. 2.16). Purines have two
fused rings, while pyrimidines have only one. The rings
contain both carbon and nitrogen atoms and form planar
2/26/2009 6:48:35 PM

A
Purines
NH2
N
H
H
N
C
N
NH2
H Guanine (G)
Pyrimidines
H
H
H
N
H
H Adenine (A)
C
C
C
C
N
C
C
C
N
C
H
H
C
C
N
C
Cytosine
Thymine
Uracil
Fig. 2.16 Structure of purine and pyrimidine bases.
Table 2.14 Nucleoside phosphates*

Base
Nucleoside
Nucleoside
monophosphates
Nucleoside
diphosphates
Nucleoside
triphosphates
Cyclic
nucleotides
Adenine
Adenosine
AMP
ADP
ATP
cAMP
Guanine
Guanosine
GMP
GDP
GTP
cGMP
Cytosine
Cytidine
CMP
CDP
CTP
Uracil
Uridine
UMP
UDP
UTP
Thymine
Thymidine
TMP
TDP
TTP
*The nucleosides A, G, C and U can contain ribose or deoxyribose, but AMP infers the ribose form, which strictly is called rAMP, the deoxyribose
form being called dAMP. The thymidine nucleotides are only produced as deoxynucleotides and are usually written without the d (e.g. TTP).
molecules. The nitrogen atoms are uncharged at neutral pH,

which means that the bases are non-polar and hydrophobic.
The major purines are adenine and guanine and the major
pyrimidines are cytosine, thymine and uracil. The coenzyme
NAD (nicotinamide adenine dinucleotide) contains a pyridine
base, nicotinamide.
Nomenclature of nucleotides
There are two different ways of naming nucleotides, which
can cause confusion. Nucleosides are nucleotides without
the phosphate; that is, they have a pentose sugar and a base.
Nucleotides with one phosphate can be called mononucle
otides or nucleoside monophosphates and so on. Many of
the different nucleoside phosphates can be found in cells
and have a variety of functions (Table 2.14).
Nucleoside phosphates
Adenosine triphosphate
The most important of the nucleoside phosphates is ad
enosine triphosphate or ATP (Fig. 2.17). The covalent
phosphoanhydride bonds which attach the second () and
third phosphate () groups to the mononucleotide, adenosine monophosphate (AMP), can be broken to release energy
for other reactions, about 7.3kcal/mol each. This is under
standard laboratory conditions, although in the conditions
ch02.indd 40
within the cell, energy released may be as high as 50kJ

(12kcal). These bonds are usually shown as ~ indicating, that
they are high energy bonds. They can be formed in two ways,
either by the transfer of a phosphate group from another
phosphorylated compound, called substrate-level phosphorylation, or by a process called oxidative phosphorylation, where electrons are transferred between a number of
intermediates in order to power the enzyme ATP synthase.
Using these mechanisms, the metabolism of a single
molecule of glucose can generate 3638 molecules of ATP
(from ADP). This is an energy transfer of about 1130KJ
(270kcal/mol) of glucose. This energy can then be used to
power other cellular processes.
Other nucleoside phosphates

Many of the nucleoside phosphates have been identified as
signalling molecules, both outside and inside cells. ATP has
been identified as a neurotransmitter at nerve terminals of
the autonomic nervous system (see Chapters 4 and 8), as
have ADP and AMP. Intracellular ATP can bind to a number
of enzymes and receptors to regulate their action.
The cyclic nucleotides, cyclic AMP (cAMP) and cyclic
guanosine monophosphate (cGMP), are important intracel
lular messengers called second messengers, involved in
signal transduction between membrane receptors and intra
2/26/2009 6:48:35 PM
Adenine
Adenosine p
NH2
C
C
Triphosphate
O
O
H
O
O
O
O
Phosphoanhydride bonds
CH2
C
H
O
H
OH
Ribose
C
H
OH
Fig. 2.17 Structure of adenosine triphosphate (ATP). The symbol ~ denotes the so-called high energy bonds.
Information box 2.7 Anti-cancer drugs may act

by inhibiting nucleotide
synthesis
One of the ways in which cancer is treated is to prevent the

rapid proliferation of the cancer cells by blocking cell
division. As the majority of normal cells will not be dividing
they will not be affected by drugs which block cell division.
Many of the side effects of anti-cancer drugs can be
accounted for by their effect on cells that are normally
rapidly dividing, such as hair and intestinal cells.
A commonly used anti-cancer drug methotrexate, acts by
inhibiting an enzyme involved in the production of thymidine
nucleotides. It is an analogue of dihydrofolate, which
acts as a competitive inhibitor of the enzyme DHFR
(dihydrofolate reductase). Other cytotoxic agents act by
inhibiting other points in the production of nucleic acids.
See also Chapter 4.
cellular proteins. The coenzymes NAD, nicotinamide adenine

dinucleotide phosphate (NADP), flavin adenine dinucleotide
(FAD) and coenzyme A are all derived from nucleotides.
Sources of nucleotides
The nucleotides required to make DNA and RNA can be
synthesised de novo by all cells. The first step involves the
synthesis of ribose, as ribose 5-phosphate, which is then
converted to a precursor for all the nucleosides, PRPP (5phosphoribosyl pyrophosphate). This precursor is then used
to produce either rUMP (see Table 2.14) from which the other
pyrimidines are produced or rIMP (inosine monophosphate),
which is the precursor for the purines. The nucleoside mono
phosphates are then phosphorylated by nucleoside kinases
to generate the bi- and triphosphates.
Ribonucleotides are required continually by all cells but
dividing cells also require deoxyribonucleotides in order to
make new DNA. These are produced from the corresponding
ribonucleoside diphosphates. The deoxyribonucleotide TMP
is produced from dUMP by a series of reactions requiring the
vitamin folic acid.
Recycling of nucleotides
The synthesis of nucleotides de novo requires a large input
of energy and there are mechanisms by which free bases,
either salvaged from endogenous sources or from nucle
otides present in the diet, can be combined with PRPP to
ch02.indd 41
produce new nucleotides. This recycling is important in some

cells, particularly in T lymphocytes. Also many obligate para
sites, such as viruses, do not have the enzymes required to
manufacture nucleotides and are completely dependent on
their hosts for a supply of nucleotides.
Deoxyribonucleic acid and

ribonucleic acids
Deoxyribonucleic acid
Only a single type of nucleic acid is made from the deoxy
ribonucleotides, deoxyribonucleic acid, or DNA, and this
is found in all cells capable of dividing.
Most of the DNA within a cell is found in the nucleus, from
which derives the name nucleic acids. It is absent from red
blood cells, which do not have a nucleus. It is also found in
the cytoplasm of prokaryotic cells (bacteria) and some viruses
(DNA viruses). A small amount of DNA is also found inside
intracellular structures called mitochondria. DNA occurs as
a double-stranded molecule, which has a characteristic
double-helical structure. This structure was first proposed by
James Watson and Francis Crick in 1953, after seeing the
X-ray crystallography data obtained by Rosalind Franklin and
Maurice Wilkins.
Ribonucleic acids
Nucleic acids made from ribonucleotides are more diverse.
Three types of ribonucleic acid (RNA) are found in eukary
otic cells. These are:
Messenger RNA (mRNA): which is copied from DNA
and used as the template for the synthesis of proteins
Transfer RNA (tRNA): small molecules which transfer
amino acids to the site of protein synthesis, the
ribosome
Ribosomal RNA (rRNA): ribosomes consist of a
combination of RNA and protein in two subunits. They
are involved in binding mRNA and tRNA during protein
synthesis.

Details of how these molecules are involved in the manu

facture of proteins are covered below. RNAs are found in all
cells (except red blood cells), including prokaryotes, and in
some viruses (RNA viruses). All viruses have either DNA or
RNA as their genetic material.
2/26/2009 6:48:35 PM

Clinical box 2.6 Excess purines can cause disease
In humans, excess purines are broken down to uric acid. If
large amounts of uric acid are produced and not removed by
the kidneys, uric acid crystals are deposited in joints and soft
tissues, resulting in gout. One treatment of gout blocks the
enzyme, xanthine oxidase, which catalyses the last steps in the
production of uric acid. The intermediate breakdown products
are more water soluble than uric acid and can be excreted.
Deficiencies in one of the enzymes involved in the recycling
pathways leads to increased purine synthesis and a
concomitant increase in uric acid, leading to a condition known
as LeschNyhan syndrome. One of the features of LeschNyhan
syndrome is gout. Patients may also go on to develop arthritis
and severe mental disorders.
Constituent bases of DNA and RNA

Each nucleic acid contains two purines and two pyrimi
dines. The purines adenine (A) and guanine (G) are found in
both DNA and RNA. However, DNA and RNA differ in one
of their pyrimidines. DNA contains the pyrimidines cytosine
(C) and thymine (T), but in RNA thymine is replaced by
uracil (U).
The nucleic acid chains are formed by the sequential link

age of nucleoside triphosphates by phosphodiester bonds
between the sugar and phosphate groups (Fig. 2.18), a
reaction catalysed by the enzymes DNA polymerase and
RNA polymerase. The free hydroxyl group of carbon-3 of
the sugar is linked to the triphosphate on the carbon-5 on
the next nucleotide, with the loss of a pyrophosphate (PPi).
The chain grows in the direction 53. This forms a sugarphosphate chain with bases attached to the other side of
each sugar (carbon-1). At physiological pH the phosphate
groups are anions, so the nucleic acids have a negative
charge.
5' end
Base
CH2
Phosphodiester
bond H
Sugar
O
P
Base
O
CH2
Sugar
H
O
Phosphate O
Base
O
CH2
Primary structure
The primary structure of DNA and RNA consists of chains of
nucleotides held together by nucleotide bonds. DNA mol
ecules are very large. The genetic material in eukaryotes is
organised in units called chromosomes, each of which con
tains a single DNA molecule. In humans, the largest of these
is nearly 10cm long when fully extended and has 23 108
nucleotides in each of its two strands.
RNA molecules vary enormously in size. Each mRNA
molecule is used to code for a protein. Therefore the length
of the mRNA is related to the size of the protein. Due to the
way in which the coding occurs, the mRNA for a protein has
at least three times more nucleotides than the number of
amino acids in the protein and may have many more. Their
sizes range from 500 to 6000 nucleotides. About 50 different
tRNA molecules are produced in animal cells, each between
70 and 90 nucleotides long.
OH
Fig. 2.18 Nucleotide bond formation.
3' end
The secondary structure of DNA is relatively simple, but it

nevertheless gives clear indications of how the molecule can
function as a source of information and also how it can be
duplicated. It is dependent on interactions called base
pairing between the bases that project from the sides of the
sugarphosphate backbone of each of the two strands. The
two strands of the DNA molecule run in opposite directions
from one another; they are anti-parallel, with one 35 chain
and one 53 chain.
Base pairing
Hydrogen bonds form between pairs of bases projecting
from each strand, each base pair consisting of a purine and
a pyrimidine. The bases can only form specific pairs, depend
ing on the number of hydrogen bonds formed between them
(Fig. 2.19). Pairs consisting of adenine and thymine can
form two hydrogen bonds, and pairs between cytosine and
guanine have three hydrogen bonds. The two types of pairs
are the same width, which means the double strands are
held at the same distance apart throughout their length, with
the two carbons from the sugarphosphate backbone being
1.08nm apart.
The planar base pairs are thus stacked one on top of
another along the core of the DNA, where hydrophobic inter
actions occur between the non-polar base pairs. The nega
tively charged phosphate groups and the sugar backbone
are on the outside.
The entire DNA molecule is extremely stable because of
the large number of hydrogen bonds between the bases and
the hydrophobic interactions between the base pairs.
Helical structure of DNA

Sugar
ch02.indd 42
While the subunits which make up DNA and RNA are fairly
similar, there are great structural differences between the
macromolecules themselves.
Secondary structure of DNA
Nucleotide bonds
Phosphate O
Nucleic acid structure
The double-stranded DNA molecule forms a regular helical

structure with 10 base pairs (bp) for each turn, advancing
3.4nm (0.34nm per base pair). The geometry of the back
bone favours a right-handed helix, which is the normal struc
ture. This type of DNA is called the B form. The two helical
grooves along the outside of the molecule have different
widths, called the major and minor grooves. The DNA mol
2/26/2009 6:48:36 PM
Two hydrogen
bonds
C
C
N
N
N
H
C
C
N
T loop
CH3
C
N
Acceptor 3
stem
C1
of sugar
Anticodon
(3 bases)
C
H
Anticodon
loop
C1
of sugar
D loop
Three hydrogen
bonds
Fig. 2.20 Generalised structure of tRNA.
H
N
H
O
N
C
H
C
N
C
C
C
C
10 nm
C1
of sugar
B
30 nm
Coiled
nucleosomes
Fig. 2.19 Base pairing between A-T (or U) and C-G.

ecule is stable as well as being quite flexible as there are no
hydrogen bonds between nucleotides above or below which
would make the double helix more rigid. While most DNA
exists in the B form, other forms of DNA are known to exist
which may be involved in interactions between DNA and
proteins or different regions of DNA.
Complementary strands of DNA

The two strands of the DNA are complementary. Because
of the constraints of base pairing (A with T and G with C) a
particular sequence on one strand must be reflected by a
specific sequence on the other. For example the sequence
TGCT in one strand would be reflected by the sequence
ACGA in the other. In this complementarity lies the key to the
way in which the sequence of bases can act as a template
for the production of identical copies.
Secondary structure of RNA

RNA molecules are single-stranded and many have a welldefined secondary structure. Stem-loops are loops of RNA
that have a base-paired sequence at the beginning and end,
or hairpins (which are shorter loops that are almost all paired).
These can form between regions of the single-stranded RNA
by base pairing between complementary regions.
The small tRNAs all have a similar structure (Fig. 2.20).
They form a cloverleaf pattern with four stem-loops, each of
which forms a short double helix. rRNA forms a complex
secondary structure containing many stem-loops. This
pattern seems to be common among a wide variety of organ
isms, suggesting a common evolutionary origin.
Tertiary structure of RNA

Only some RNAs have a unique folding pattern and, as previ
ously described for proteins, the primary structure is all that
is required to reassemble the molecule if it is denatured. The
ch02.indd 43
DNA
C1
of sugar
Histones
C
300 nm
Chromatin
Protein
scaffold
D
700 nm
Fig. 2.21 Tertiary structure of DNA.

tertiary structure of tRNAs involves hydrogen bonding, which
holds two of the loops parallel to the rest of the molecule in
an L-shaped molecule, with a site for binding to mRNA at
one end and a site at the other end where the amino acid
can be attached.
Tertiary structure of DNA

In a human cell there are about 3 109 base pairs of DNA
that must be packed into the nucleus. The largest chromosome has over 200 million base pairs, which, if extended
completely, would give a DNA molecule that would be much
larger than the cell. In addition, the packing must allow this
DNA to be easily accessible by uncoiling, in order to copy
segments.
Chromatin and nucleosomes

In the chromosome, DNA is combined with some RNA and
about the same mass of protein, producing a structure called
chromatin. Chromatin consists of a series of units called
nucleosomes (Fig. 2.21). Each nucleosome is composed of
globular proteins called histones, around which are wrapped
two loops of DNA 140bp long, with a piece of linker DNA
between each nucleosome about 80bp long (Fig. 2.21A).
2/26/2009 6:48:36 PM
Information box 2.8 The Human Genome Project

has sequenced the entire
human genome
A project to map and sequence the entire human genome

of approximately 3 billion base pairs was started in 1990
and the first stage, described as a working draft, was
declared complete on 26 June 2000, by two groups, the
Human Genome Project led by Francis Collins, and Celera
Genomics led by Craig Venter. The groups used different
techniques to produce the draft, which was fully completed
by 2003. Surprisingly the total number of genes in the
human genome turned out to be much lower than expected
(2000040000) although many genes produce more than
one gene product through alternative splicing. Much effort
is now concerned with identifying the role of these genes
and it is expected that this will lead to further advances in
medicine.
This produces a 10nm width fibre which is then coiled into

a fibre 30nm wide with 6 nucleosomes per turn (Fig. 2.21B).
The coils are then formed into a supercoiled structure
300nm in diameter (Fig. 2.21C). This multiple folding reduces
the length of the DNA molecule by about 8000-fold. When
cells are about to divide the chromatin is concentrated still
further and the DNA molecules become visible under a light
microscope as chromosomes (Fig. 2.21D). How DNA uncoils
in order to be copied is an obvious topological problem.
Human cells have 23 pairs of chromosomes, each with a
specific size and shape.
Mitochondrial DNA
The tertiary structure of mitochondrial DNA is very different
from nuclear DNA. It is a circular molecule, which is
similar to the DNA in bacteria. This is used as evidence that
mitochondria originated as free-living bacteria, before forming
a symbiotic relationship with what later became eukaryotic
cells.
BASIC CELL STRUCTURE

The defining difference between prokaryotic and eukaryotic
cells is the presence, in all eukaryotic cells, of a nucleus and
other intracellular structures. In the same way that macro
molecules contained within cells have similarities across a
wide range of organisms, so there are common structures
found in cells of different types. However, the specific struc
ture of a particular cell varies enormously, both between cells
of different organisms and between cells within a multicellular
organism. While there are many multicellular organisms, such
as sponges, which consist of aggregates of largely similar
cells, most multicellular organisms have cells with different
specialised functions. Furthermore, animals and plants have
significant differences in their organelles and cell
membranes.
HISTOLOGY
Cellular structures can be identified using either light micro
scopy or electron microscopy (EM). The study of tissue
structure by microscopy is termed histology. Naturally, the
thin layers of cells or tissues on a slide are transparent and
ch02.indd 44
featureless, so both types of microscopy require that the cell

or components within the cell are made opaque in some
way, which usually kills the tissue. Specialised forms of light
microscopy, such as phase contrast, can be used to observe
living specimens, although their resolution is fairly poor. The
light microscope can resolve objects which are about 0.2m
apart with magnification up to about 1000 times.
Histochemistry
Cells can be treated by a wide range of methods so that they
become visible under the light microscope. These methods
rely on specific chemical reactions between the chemicals
being applied and the molecules making up or contained
within the cellular organelles. One of the most useful stains
is a combination called haematoxylin and eosin (H and E).
Haematoxylin is a basic dye that stains acidic components
in the cell blue. Eosin is an acidic dye that stains basic com
ponents pink. In most cells, H and E stains the nucleus,
containing the acidic DNA and RNA, blue, and the cytoplasm,
most of whose proteins are basic, pink.
Other staining methods have been developed in order to
label specific tissues; for example enzyme histochemistry
can identify cells or parts of cells containing a particular
enzyme. The recent development of methods using immuno
logical markers, a technique referred to as immunohisto
chemistry, enables identification of specific proteins or other
macromolecules within cells.
Electron microscopy
The detailed ultrastructure of cells can be revealed only by
electron microscopy, where the image is formed by bom
barding the specimen with electrons. This can resolve objects
which are 1nm apart, with a maximum magnification of
about 100000-fold. There are two types of EM:

Scanning EM used to image the surface of objects.

Transmission EM which uses thin sections of tissue
treated with heavy metals that bind to cell components
in varying amounts and reflect electrons. Those
components with a large amount of bound metal reflect
many electrons, are electron dense, and look dark in
EM, and vice versa.
All EM images are black and white only, although false

colour processing can be used to highlight differences in
grey scale.
Artefacts
Care must be taken in interpreting images from both types
of microscopy. Artefacts can be introduced during the many
steps involved in the preparation of a specimen, and inter
preting two-dimensional images as three-dimensional (3D)
objects takes a lot of thought. However, computer software
nowadays assists in reconstructing 3D structures from
sequential sections.
ORGANELLES, STRUCTURE
AND FUNCTION
Organelles are distinctive structures found within cells.
Many are enclosed by one or more membranes, but there
are a few structures which are not (called non-membranous
organelles) (Fig. 2.22). Organelles enable cells to compart
mentalise biochemical processes into different regions of the
cell. Enzymes needed to carry out a particular reaction can
2/26/2009 6:48:37 PM
Basic cell structure 45

Rough endoplastic
reticulum
Nucleolus
Mitochondria
Nucleus
Information box 2.9 Gram staining of bacterial

cell walls
Gram staining was invented by Hans Christian Gram in

1884 and is used to identify different types of bacteria. The
method consists of soaking the sample of the bacterial
culture firstly in a violet dye (Gentian violet), followed by
treatment with iodine. The slide is then washed with alcohol
and counter-stained with a red dye, safranine.
Gram-positive bacteria are coloured blue-black and
Gram-negative bacteria are stained red. The different
colours obtained are due to the different amounts and
accessibility of the peptidoglycan in the cell walls of
bacteria. Gram-positive bacteria have a thick layer of
peptidoglycan surrounding their plasma membranes. This
takes up the violet dye and appears blue to purple.
Gram-negative bacteria have less peptidoglycan, which is
surrounded by a second outer cell membrane.
Mycobacteria, such as those causing tuberculosis, have
cell walls that have a very different composition and are not
stained by the Gram stain.
Golgi
Lysosomes
Smooth endoplasmic
reticulum
Fig. 2.22 An idealised eukaryotic cell showing the

major organelles. Not all organelles are present in all cells.
be put in the same compartment as the substrates for the

reaction, and the products can be passed onto the next
enzyme in the chain by placing it in the same compartment.
Conditions can be maintained within the restricted volume of
organelles that would be harmful to the cell overall.
Cell membranes
All cells, both prokaryotic and eukaryotic, are surrounded by
a plasma membrane, also called the cell membrane. It
maintains the integrity of the cell and separates the fluids
inside the cell from those on the outside and is typically
about 9nm thick. Under the electron microscope it is seen
to consist of two outer dark layers, with a clearer inner layer.
The plasma membrane regulates the movement of molecules
in and out of the cell. The major component of all membranes
is lipid, which on its own would make the cell impermeable
to all hydrophilic molecules. However, it also contains pro
teins that allow the movement of hydrophilic molecules
through the membrane. The outside of the plasma mem
brane is also studded with carbohydrates, attached to the
proteins and the lipids, which have roles in cell-to-cell inter
actions and cell recognition. The membranes which surround
the membrane-bound organelles also have a banded,
double-layered appearance in electron micrographs but are
thinner (about 6nm).
Plant and bacterial cell walls

Plant cells have a rigid cell wall outside their plasma mem
brane, which gives the plant cell strength. Plant cell walls
contain cellulose and sometimes lignin and suberin, which
form wood and cork, respectively. Most bacteria also have
cell walls. They often contain peptidoglycan, a polymer of
amino sugars cross-linked with short peptide chains. The
presence and concentration of this macromolecule is
used as a way to categorise bacteria. Peptidoglycan is not
ch02.indd 45
found in eukaryotic cells and many antibiotics, including the

penicillins, inhibit bacterial cell growth by inhibiting its
synthesis.
Cytoplasm
Between the plasma membrane and the nucleus is the
cytoplasm, which contains all the other cellular elements
suspended in the cytosol. This is a viscous aqueous solution
that contains salts, proteins, sugars and the countless other
molecules that are involved in the metabolic pathways. It is
criss-crossed by the fibrous cytoskeleton. It has been esti
mated that there are about 300mg/mL of protein and RNA
in the cytosol, which makes it very crowded indeed.
Nucleus
The most prominent organelle in most eukaryotic cells is the
nucleus, which is often at the centre of the cell, although
some cells, such as skeletal muscle cells, have their nuclei
displaced to one side. Some cells have more than one
nucleus, whereas mammalian red blood cells lose their
nucleus as they mature (see Ch. 12). The nucleus is the only
organelle that can be seen under the light microscope with
out staining.
The nucleus contains the chromosomes in the form of
chromatin, which is not normally visible as an organised
structure. Actively transcribed chromatin, called euchroma
tin, is diffuse. This is where mRNA and tRNA are being tran
scribed. More condensed chromatin, called heterochromatin,
is inactive.
Inside the nucleus are one or more dark-staining spherical
areas called nucleoli. These are areas of the nucleus where
ribosomes are assembled. These areas contain large amounts
of RNA and proteins, and in cells which are actively secreting
large amounts of protein they can be very large. Surrounding
the chromatin and the nucleoli is a gel-like suspension called
the nucleoplasm.
The nucleus is surrounded by a double layer of mem
brane, the nuclear envelope. The inner membrane is smooth,
whereas the outer membrane may be continuous with the
2/26/2009 6:48:37 PM
Information box 2.10 Barr bodies represent

inactive X chromosomes
In all female cells, there are two X chromosomes. However,

in all somatic cells (that are not oocytes), one of these X
chromosomes is inactivated at random. In about 30% of
nuclei this can be seen as a condensed mass of
heterochromatin called a Barr body, or X chromatin. In
white blood cells called neutrophils this appears as a small
drumstick. If the cell contains more than two X
chromosomes, such as in triple X syndrome (XXX), then the
extra chromosomes will be seen as extra Barr bodies.
endoplasmic reticulum and the outer face may be covered

with ribosomes. The nuclear envelope is studded with pores.
These are about 9nm in diameter and are surrounded by an
octet of protein complexes in each membrane, forming a
channel through which molecules of protein and RNA can
move.
Endoplasmic reticulum
Extending from the nucleus and into the cytoplasm are a
series of interconnected flattened tubules and sacs called
the endoplasmic reticulum (ER). This network of mem
branes extending into the cytoplasm, which is continuous
with the outer nuclear membrane, encloses cavities called
cisternae. There are two types of ER within the cell, one
called rough ER, whose membranes are covered with dense
granules, and smooth ER, which does not have these
granules.
Ribosomes and rough ER

The granules which are studded all over the rough ER are
called ribosomes. They are made from two subunits, each
made up of a combination of RNA and protein which is
assembled in the nucleolus. The large subunit contains a
large RNA molecule of 4800 nucleotides with two smaller
RNAs of 160 and 120 nucleotides, all associated with about
50 proteins. The small subunit has a single RNA of 1900
nucleotides and about 33 proteins.
Ribosomes are involved in the synthesis of proteins,
which may be destined for insertion into membranes, used
in lysosomes, or exported from the cell. Rough ER is very
extensive in cells that have high levels of protein secretion.
Many of these proteins are then processed by enzymes
within the ER, which can add sugars and proteins to them.
Small vesicles bud off from the ER and are seen to combine
with the Golgi apparatus (see below).
Ribosomes are also found in the cytoplasm, unattached
to the ER. These free ribosomes are involved in the manu
facture of cytoplasmic and nuclear proteins. Mitochondria
also contain ribosomes, whose rRNA is coded for by mito
chondrial DNA.
Smooth ER
Smooth ER is continuous with rough ER, but is generally
more tubular in nature. Smooth ER manufactures membrane
phospholipids and also makes cholesterol and steroid hor
mones. Cells, such as those in the adrenal cortex, the testes
and the ovaries, which secrete large amounts of steroid hor
mones, have very extensive smooth ER. In the liver, cells
ch02.indd 46
called hepatocytes have rough ER, which is involved in the

production of plasma proteins, such as albumin. However, a
large proportion of the ER in these cells consists of smooth
ER. This smooth ER is associated with granules of glycogen,
which can be broken down to maintain blood glucose levels.
The enzymes in the smooth ER in the liver are involved in the
detoxification of a wide range of lipid-soluble molecules,
including carcinogens and drugs, transforming them into
polar metabolites that can then be excreted by the kidney
(see Ch. 14). In two types of muscle cells, cardiac and skel
etal muscle, a specialised smooth ER, called sarcoplasmic
reticulum, has a critical role in the sequestration and release
of calcium ions that are required for the activation of muscle
contraction.
Golgi apparatus
The Golgi apparatus, named after Camillo Golgi, who first
described it in 1898, appears as a number of flattened
membrane-bound sacs called cisternae, stacked on top of
each other and surrounded by large numbers of small spheri
cal membrane-bound organelles called vesicles. It is involved
in the concentration, modification (mainly glycosylation and
phosphorylation) and packaging of proteins produced in
the rough ER. The Golgi apparatus has a definite polarity.
Small vesicles, called transport vesicles, derived from the ER
and containing proteins and lipids, fuse onto the cis Golgi.
After processing through the medial Golgi, where they are
glycosylated, vesicles containing protein bud off from the
trans Golgi, where they are sorted for a number of different
destinations including secretory vesicles and lysosomes. The
proteins move through the Golgi stack by being shuttled in
small transport vesicles from cisterna to cisterna.
Secretory vesicles
Proteins destined to be exported from the cell are contained
in vesicles derived from the trans Golgi. Mature vesicles are
called secretory granules, and in some cells (for example,
in the exocrine pancreas) these are some of the most
prominent structures, filling a large proportion of the cyto
plasm. During maturation the contents of the vesicles are
concentrated and the proteins can be modified; for example
modification of the insulin molecule, which removes the
C-peptide, occurs in the secretory granules (see Clinical
Box 2.1).
Secretion
Proteins are released into the extracellular space by a process
called exocytosis, which involves the fusion of the vesicles
membrane with the plasma membrane. Release of the pro
teins may be continuous, or vesicles may be stored. Continu
ous secretion, called constitutive secretion, occurs in some
immune cells called plasma cells, which continually secrete
immunoglobulins.
Most secretion occurs in response to a signal, often a rise
in intracellular calcium levels, and this regulated secretion
releases stored secretory granules. Many neurons release
more than one type of chemical and it is possible to identify
different populations of vesicles in nerve terminals. The dif
ferent chemicals, which often consist of a small non-protein
and a protein, are released under different patterns of stimu
lation (see Ch. 8).
2/26/2009 6:48:37 PM
Information box 2.11 Enzyme histochemistry can

be used to identify
lysosomes
Despite their variable and relatively unremarkable structure,

lysosomes can be easily identified using a staining
technique called enzyme histochemistry, as they contain
acid phosphatases. The cells are incubated with a
substrate for the acid phosphatase. The reaction between
enzyme and substrate releases phosphate ions. These ions
react with lead ions to form the insoluble product lead
phosphate, which is deposited at the location of the acid
phosphatase. This is then treated with ammonium sulphide
to give a black precipitate of lead sulphide, clearly visible
with a light or electron microscope.
Lysosomes and proteasomes

One of the destinations for vesicles derived from the Golgi
are the lysosomes. These are spherical or oval organelles
with a variety of sizes up to 400nm diameter. They contain
about 40 different acid hydrolase enzymes which can digest
most biological molecules. As well as vesicles from trans
Golgi, other membrane-bound structures fuse with lyso
somes, where their contents can be digested. These include
particles such as bacteria and viruses that have been
retrieved from the extracellular medium by two processes
called endocytosis and phagocytosis. The lysosome is also
used to destroy aged cell organelles that are first engulfed
by the ER, forming a vesicle that is passed to lysosomes for
destruction.
Acid hydrolases
The fluid inside lysosomes is much more acidic, at about
pH4.8, than the normal pH of about 7.07.3. This is the
optimal pH for the activity of the acid hydrolase enzymes
present in the lysosomes. The low pH denatures many
macromolecules, which aids their degradation. However, for
tunately, the acid hydrolase enzymes are relatively inactive at
the pH of the cytoplasm, so if a single lysosome is ruptured
little digestion of cytoplasmic contents occurs. Hydrogen
ions will tend to leak out of the lysosome so the pH gradient
between the cytoplasm and the interior of the lysosome is
maintained by special protein pumps in the lysosomal mem
brane that pump hydrogen ions into the lysosome. Other
pumps transport the products of the lysosomal digestion,
such as amino acids and sugars, into the cytoplasm where
they can be recycled.
The lysosomal membrane has an important role in com
partmentalising the degradative enzymes because, if many
lysosomes were ruptured, the cell could self-digest. This
process is known as autolysis and can occur physiologically,
such as, for example, during the breakdown of the uterine
lining during menstruation.
Proteasomes
While membrane components and aged organelles are
degraded in lysosomes, another pathway exists to degrade
cytosolic proteins. Proteasomes are large complexes of
proteins arranged in four rings around a central core. Pro
teins destined for destruction are first tagged with multiple
copies of a protein called ubiquitin by specific enzymes. This
acts as a signal to direct the protein into the core of the
proteasome where it is broken into small peptides. These
ch02.indd 47
Clinical box 2.7 TaySachs disease is one of a

number of lysosomal storage
diseases
The absence of a particular enzyme in the breakdown pathways
inside lysosomes leads to the accumulation of the substrate for
that enzyme. In TaySachs disease, the absence of the
enzyme, hexosaminidase leads to the accumulation of a
membrane glycolipid, GM2, which is particularly common in
nerve cells. The disease, which is inherited, causes mental
retardation and central nervous system disorders, with death
occurring by about the age of 5.
are then further digested in the cytoplasm by peptidases.

Only proteins tagged with ubiquitin are able to enter the
proteasomes, ensuring that normal cytosolic proteins are
protected.
Many proteins are continually being turned over and each
protein has its own rate of degradation. Some enzymes are
replaced rapidly but others such as collagen have an almost
imperceptible turnover rate. In starvation, proteins, particu
larly skeletal muscle proteins, are broken down to provide
precursors for glucose synthesis.
Peroxisomes
Peroxisomes are small vesicles, also called microbodies,
which superficially resemble lysosomes, but contain com
pletely different enzymes that are transported into the peroxi
some from the cytosol. Some of these enzymes are oxidases
and catalases:

The oxidases break down a variety of substrates, such

as fatty acids, with the production of hydrogen peroxide
(H2O2). Although potentially highly toxic to cells, H2O2 is
produced inside some cells of the immune system in
order to kill bacteria.
The catalase enzymes use H2O2 to oxidise other
potentially toxic compounds, or break down H2O2 to
water.
Other peroxidase enzymes are also involved in the bio
synthesis of cholesterol and other membrane components.
Peroxisomes are particularly abundant in the hepatocytes of
the liver.
Mitochondria
All the organelles described so far are involved in the produc
tion, processing, targeting and degradation of the proteins
and other molecules made within the cell. However, most of
these processes require energy in the form of ATP. Mitochondria are the organelles where ATP is generated, and so
they are particularly plentiful in cells which consume large
amounts of energy.
Structure of mitochondria
Mitochondria vary in size and shape but are generally
sausage-shaped and about 1m wide and 7m long. They
have a complex internal structure with two layers of mem
branes. The outer membrane is smooth and the same thick
ness as in other organelles. It is highly permeable to small
molecules, due to the presence of a pore-forming protein
called porin. The intermembrane space between the outer
2/26/2009 6:48:37 PM

and inner membrane is usually narrow. The inner membrane
is studded with many proteins that are involved in oxidative
phosphorylation, and the subsequent generation of ATP. This
membrane is thinner and has multiple folds projecting
inwards, called cristae, producing a very large surface area
for ATP production. The innermost region is called the matrix.
It is here that pyruvate, derived from the metabolism of
glucose in the cytosol, and fatty acids are metabolised to
provide the substrates for oxidative phosphorylation.
Mitochondrial DNA
The mitochondria are unique among the organelles, in that
they contain their own DNA and the mechanism to make
proteins. The matrix contains mitochondrial DNA (mtDNA),
which codes for some of the molecules needed by the mito
chondrion, including its own rRNA and tRNA. However, many
of the proteins required by mitochondria are manufactured in
the cytoplasm and have to be imported into the mitochon
drial matrix. Receptors on the surface of mitochondria act to
transfer these proteins required in the matrix through the
outer and inner membranes.
Evolutionary origin of mitochondria

Bacterial cells do not contain mitochondria but mitochondria
themselves have many similarities with a group of bacteria
called purple bacteria. This has led to the hypothesis called
the endosymbiotic theory: that mitochondria and other
intracellular organelles of eukaryotes could have arisen when
one type of bacteria was engulfed by another, without digest
ing it. The two bacteria could have become dependent on
one another until they could no longer exist separately.
Another piece of evidence for the bacterial origin of mito
chondria is that the genetic code which translates the bases
of the mitochondrial DNA molecule into amino acids is slightly
different from that used by the rest of the cell, and can vary
both between different mammals and other organisms.
Inheritance of mitochondrial DNA

Mitochondria are passed from parent to child during repro
duction, but because the volume of the cytoplasm contrib
uted by the sperm is very small compared to that of the
ovum, virtually all the mitochondria (estimated at 99.99%)
are inherited from the mother. mtDNA is therefore passed,
virtually unchanged, down the maternal line and geneticists
interested in the evolution of humans have utilised mtDNA
lineages to examine population movements.
Plant mitochondria
In plants, the equivalent organelle is the chloroplast, which
generates glucose from light, water and CO2 by photosyn
thesis. This reaction not only provides all the food we eat,
but also produces oxygen as a waste product, without
which we could not live.
Non-membranous organelles
Non-membranous organelles are cellular structures not
bound by membranes. These include ribosomes and lipid
droplets, which are composed of triacylglycerols, seen in
fat-storing cells called adipocytes. In liver hepatocytes, large
numbers of glycogen granules can be seen in the cytoplasm
either as single particles or as aggregates called rosettes.
Cytoskeleton
All cells have an internal skeleton of protein fibres called the
cytoskeleton (Table 2.9). These are made up of:

Microfilaments
Intermediate filaments
Microtubules.

Microfilaments
Microfilaments are composed of twisted double strands of
actin (Fig. 2.23A) and are the thinnest of the cytoskeletal
fibres at 7nm in diameter. They enable cells to move in an
amoeboid fashion and change their shape. They form a mesh
of fibres below the plasma membrane attached to proteins
embedded in the membrane, which themselves connect with
A Microfilaments
7 nm
Actin molecule
B Intermediate filaments
Head
Rod
Tail
IF monomer
helix
Parallel dimer forms
10 nm
C Microtubules
Tubulin dimer
13 nm
Information box 2.12 Mutations in mitochondrial

DNA are inherited
maternally
Any mutations that occur in mtDNA are passed from a

mother to all her children. Because many of these
mutations occur in the enzymes responsible for energy
metabolism and impair ATP production they often result in
an increase in anaerobic metabolism, with the production of
lactic acid. This may cause cell death in tissues such as
skeletal and cardiac muscle which are heavily dependent
on ATP, causing myopathies.
Singlet
Doublet
Triplet
A
B
A
B
C
Fig. 2.23 Structure of cytoskeletal elements. IF,

intermediate filament.
ch02.indd 48
2/26/2009 6:48:38 PM

molecules of the ECM. This enables a cell to fix its shape
with respect to the external environment and, if necessary,
move around in it. Cells move by extending either thin
finger-like structures, called filopodia, or thin sheets, called
lamellipodia, which contain large amounts of F-actin that is
continuously added to in the direction of movement. The
fibres are thought to be moved forward by the movement of
myosin pushing the actin fibres against the plasma mem
brane. They are attached to long actin fibres, called stress
fibres, which extend backwards. Microfilaments also form
the core of the specialised plasma membrane structures,
microvilli and stereocilium.
required in the synapse are manufactured in and near the

nucleus in the cell body and must be transported along
microtubules to the synapse. This process is called fast
axonal transport.
Transport occurs in both directions along the same
microtubule and is rapid (12m/s):
Intermediate filaments
The anterograde transport, from the - end to the + end of

each microtubule, is carried out by proteins called kinesins.
These consist of four peptide chains: two heavy chains that
bind to tubulin at their head end, and which are then coiled
in an -helix until they reach the tail end, where there are two
light chains which bind to the transported substance. The
force required to move material is provided by the heavy
chains which, like the muscle protein myosin, change their
shape on the hydrolysis of ATP.
Intermediate filaments (IFs) are made from a variety of

proteins. However, they all form filaments 10nm wide which
are extremely stable. They all form -helical dimers, which
twist around one another forming rope-like structures
(Fig. 2.23B), which give mechanical support to structures
such as the nucleus and the plasma membrane. IFs do
not change dynamically like microfilaments and micro
tubules, and so they form the most stable element of the
cytoskeleton.
There are five classes of IFs, which are characteristic of
each cell type. In epithelial cells, IFs made from keratin are
anchored to structures called desmosomes, which link cells
together, and hemidesmosomes, which link cells to the ECM.
In nerve cells, neurofilaments form the core of axons which
can stretch for long distances.
Microtubules
Microtubules are made up of two globular proteins,
- and -tubulin, which form a dimer. These proteins are
polymerised into a protofilament of alternating - and tubulin. These are then arranged to form a tube from 13
protofilaments, which is 25nm wide, called a singlet (Fig.
2.23C). Other arrangements involve adding extra protofila
ments around the singlet to form doublets and triplets. The
microtubule can be extended by the addition of further
dimers to the + end of each protofilament, the end furthest
away from the nucleus. Microtubules can be shortened
from both ends by disassembly, which, using electron micro
scopy, looks as if the tube is being frayed. The growth of
microtubules requires GTP, which is hydrolysed to GDP
when the dimer of tubulin is incorporated into the growing
protofilaments.
A number of different proteins are attached to micro
tubules, called microtubule-associated proteins (MAPs).
One type of MAP cross-links microtubules in the cytoplasm,
or binds to IFs. One of the MAPs found in nerve cells is called
tau. A model of Alzheimers disease suggests that accumula
tion and cross-linking of tau proteins can lead to the forma
tion of the neurofibrillary tangles which are found in the brains
of many people with Alzheimers.
Transport along microtubules

Inside the cell, organelles and proteins are moved to specific
locations. Microtubules are used as tracks along which
organelles can be moved. For example, in nerve cells, the
distance between the nucleus and the synapse, the point at
the end of the axon where chemicals are released, can be
several metres. The synapse does not contain the ribosomes
required for protein synthesis, and in any case the DNA is in
the distant nucleus. Proteins and membrane components
ch02.indd 49
Anterograde transport carries new material from the

cell body to the synapse.
Retrograde transport carries unwanted materials back
to the cell body for destruction by lysosomes.

Anterograde transport
Retrograde transport
Another type of motor protein, dynein, is responsible for
retrograde transport. Dyneins have a different structure from
kinesins but they are similar in that a globular head region
binds to the microtubules and moves with the consumption
of ATP. In nerve cells, microtubules do not extend all the way
to the synapse; they stop at the end of the axon, where they
are replaced by microfilaments. Materials transported along
the microtubules are taken the rest of the way along these
microfilaments, the movement being powered by a type of
myosin and ATP. Other, slower transport may involve IFs.
Microtubule structures
Three distinct structures are formed from microtubules:

Centrosomes
Cilia
Flagella.

Centrosomes
Most cells contain a centrosome which is found near the
nucleus. This is the organising centre for microtubules, which
radiate from here to the plasma membrane. The centrosome
contains two centrioles, each of which is composed of a
cylinder made up of nine microtubule triplets. This arrange
ment is called a 9 + 0 array. Centrioles are essential to the
movement of the chromosomes to the opposite ends of a
cell during cell division.
Cilia and flagella

Cilia and flagella (singular: cilium and flagellum) are mobile
projections from the plasma membrane that are also com
posed of microtubules. The arrangement is slightly different
from that found in centrioles, as the core or axoneme is
formed from nine microtubule doublets, arranged around a
doublet running down the central axis, in what is called a
9 + 2 array. Just underneath the plasma membrane, cilia and
flagella are anchored to the basal body, which is identical in
structure to the centrioles. The doublets of the cilia and
flagella merge into the triplets of the basal body. The entire
structure is covered in extensions of the plasma membrane.
Inner and outer arms attached to the nine doublets are
2/26/2009 6:48:39 PM

Clinical box 2.8 Cigarette smoke stops the
mucociliary escalator
Clinical box 2.9 Cardiac muscle is very sensitive

to changes in K+ concentration
The cilia which move mucus and trapped particles out of the
lungs are inhibited and eventually destroyed by the inhalation of
cigarette smoke. When this occurs, the only way that mucus
can be removed from the lungs is by coughing. Irritants in the
smoke cause chronic inflammation of the airways leading to
excess production of thick mucus. This narrows the airways
and traps pathogens, which can cause infection. Chronic
bronchitis, along with emphysema, are two pathologies found
in chronic obstructive pulmonary disease (COPD), a serious
respiratory disease which occurs almost exclusively in smokers.
See also Chapter 5.
Changes in the extracellular K+ concentration alter the resting

membrane potential of cardiac myocytes, which can change
their excitability. Normal extracellular K+ levels are 3.55.0mM
but if levels fall below this then the myocytes will become
hyperpolarised and cardiac excitation is reduced. If the K+
concentration rises above 5.5mM, at first the cardiac excitation
increases, which can cause arrhythmias and may lead to heart
attacks (ventricular fibrillation). However, if K+ levels increase
further, then heart block may occur. Levels of K+ above 7mM
can cause complete heart block, where the heart stops
completely (see Ch. 11).
dyneins, which move the cilia and flagella using energy

derived from ATP.
Cilia are hair-like projections up to 10m long, whereas
flagella are much longer and occur individually or in pairs.
Many cells are covered with cilia, which serve to move fluids
or particles across their surface. There may be up to 300 cilia
on each cell and they move in a coordinated manner. In the
lungs, the cells which cover the larger airways are covered
in cilia, which move mucus and particulates out of the lungs
the mucociliary escalator.
In single-celled organisms, cilia and flagella are the means
by which the organism propels itself through the medium in
which it lives. The only human cells that have flagella are
sperm. Each sperm moves by beating the flagellum that
forms its tail. Many bacteria also have flagella but these
are completely different in structure, consisting of a protein
strand that is rotated in its basal body, using energy derived
from proton gradients.
protein. Free calcium levels inside cells are very low, reflect
ing its role as an intracellular messenger.
INTRACELLULAR AND
EXTRACELLULAR FLUIDS
All cells contain fluid and are bathed in fluid, but the compo
sition of the intracellular and extracellular fluids is very differ
ent (Table 2.15). The extracellular fluid resembles a dilute
version of seawater, possibly reflecting our evolutionary
origins as single-celled marine organisms. It is rich in sodium
and chloride ions, with quite high levels of calcium but very
little protein compared with intracellular fluids. The intracellular fluid has a high concentration of potassium and a large
number of intracellular anions such as phosphates and
protein side chains, which balance the positively charged
potassium ions. It also has a relatively high concentration of
Ion gradients
While a pure lipid bilayer is virtually impermeable to ions,
there are a number of transport processes acting across cell
membranes which not only allow the dissipation of the ion
gradient but also use the energy contained within it. The
concentration gradients of Na+ and K+ across the plasma
membrane are maintained by the activity of a special trans
port protein called the Na+/K+ATPase, which transports
sodium ions out of the cell and potassium into it. Without the
activity of this pump the ion gradients would disappear and
many cellular processes would cease to function.
Resting membrane potential

The differential distribution of sodium and potassium cations,
and the fact that cells are more permeable (at rest) to potas
sium than sodium, leads to the establishment of an electrical
potential. This is called the resting membrane potential
which acts across the cell membrane. In most cells it is
of the order of -60mV (inside negative), but is higher in
excitable cells, such as neurons and muscle (see Ch. 8).
Variations in this electrical potential underlie the electrical
signalling carried out by excitable cells, which is entirely
dependent on the ion gradients of sodium and potassium.
The resting membrane potential is very sensitive to the
external potassium concentration, which must always be
closely controlled otherwise the electrical signalling will be
disrupted (Clinical box 2.9).
Osmotic pressure
Osmolarity
Table 2.15 Composition of intracellular and

extracellular fluids
Ion
Extracellular
concentration
(mM)
Na+
145
12
K+
139
Cl
116
29
12
Bicarbonate, HCO3
ch02.indd 50
Intracellular
concentration
(mM)
Phosphates
13
Protein anions
138
Ca2+
1.8
Mg2+
1.5
<0.0002
0.8
Osmosis is the movement of water by diffusion through a

semi-permeable membrane from a region of high water con
centration to a lower concentration. If two solutions of differ
ing solute concentration are separated by a semi-permeable
membrane water will tend to move from the lower solute
concentration to the higher one until the solutions have equal
concentrations of both water and solute. The solution with
the higher solute concentration is said to have a higher
osmotic potential.
Water is a polar molecule and pure phospholipid bilayers,
which are non-polar, only allow limited water movement
across them. However, in many cells, particularly in erythro
cytes and water-absorbing cells of the collecting ducts of the
kidney, there are large numbers of specific water channels,
formed by proteins called aquaporins. Using these channels
2/26/2009 6:48:39 PM
Protein synthesis and processing 51

Clinical box 2.10 Intravenous fluid must have the
right osmolarity
Clinical box 2.11 Low albumin levels in the blood

lead to oedema
When administering intravenous fluids it is important to match

not only the correct ionic concentrations, but also the correct
osmotic potential. The osmolarity of body fluids is about
285mOsm/L. If an isotonic sodium chloride solution is added to
the extracellular fluid, because the sodium and chloride remain
outside the cells, there is no osmotic movement of water into
the cells. However, if a hypertonic sodium chloride solution is
administered, water is drawn out of the cells, further increasing
the extracellular fluid volume, with an increase in the osmolarity
of both compartments. Severe hypotension is often treated by a
rapid infusion of hypertonic solutions, increasing the circulating
plasma volume. The inverse occurs with a hypotonic solution.
There is an increase in the intracellular volume as water diffuses
into the more concentrated intracellular compartment and both
compartments are enlarged, with a reduction in osmolarity.
In liver disease, the levels of albumin in the blood may be

lowered as the liver loses its ability to make enough protein.
Equally, in the kidney, if there is an increased permeability that
allows proteins to be lost in the urine (they are normally
retained), then the concentration of albumin in the blood will fall.
If this happens then the osmotic potential of the blood will be
reduced and water will tend to accumulate in the extravascular
compartment. This causes the tissues to become puffy and
they will indent when pressed. This is more evident in the lower
limbs as water accumulates there under the force of gravity.
Called pitting oedema, this is a diagnostic sign of right heart
failure, as well as of some liver and kidney disorders.
water moves freely across these plasma membranes. There

fore the internal and external media must have the same
osmotic potential (they must be isotonic), otherwise cells
would shrink or swell and burst. If cells, for example eryth
rocytes, are put in a dilute solution which has a lower osmotic
potential (it is hypotonic) than normal plasma, then water will
move into the cells and they will swell and eventually break
open (lysis). Equally, if they are put into a more concentrated
hypertonic solution then water would move out of the cells
and they would shrink. The osmotic pressure of the intracel
lular and extracellular fluids is normally the same at about
285 milliosmoles per litre (mOsm/L (see Ch. 14)).
Vascular compartment
The intravascular compartment, which contains the blood,
is separated from the extracellular fluid surrounding most
tissues by a barrier, which in the smallest blood vessels, the
capillaries, consists of endothelial cells. Although in some
tissues, such as the liver, this barrier is very permeable to
both water and solutes, in most tissues there is a significant
difference between the intravascular and extracellular fluid
composition. Although they both contain sodium as their
predominant cation, the blood also contains significant
amounts of protein, mainly in the form of albumin. This means
that blood has a higher osmotic potential than extracellular
fluid, which tends to draw water out of the interstitial spaces
between the cells. This does not mean that all the fluid
around cells moves into the blood. The blood has a signifi
cant hydrostatic pressure and this tends to push water out
of the blood and into the interstitium. While this varies around
the body, the net effect is that the movement of water into
and out of the vasculature is balanced (see Clinical box 2.11
and Ch. 11).
PROTEIN SYNTHESIS AND

PROCESSING
The central dogma of molecular biology is that, briefly stated,
DNA makes RNA makes protein and not the other way
round. That is, you cannot make protein from protein or DNA
from RNA. This holds true for all cellular organisms. Only in
some viruses, which are obligate parasites, is this rule broken
(see Information box 2.14).
ch02.indd 51
The synthesis of proteins involves two processes which

convert the base sequence of the appropriate DNA into a
polypeptide chain.
1. Transcription: the conversion of a specific section of
DNA into a complementary strand of mRNA. This
transcription occurs in the nucleus after which the
mRNA is processed and translocated to the cytoplasm
where it is attached to a ribosome.
2. Translation: the strand of mRNA is used as a template
to assemble the sequence of amino acids which make
up the protein. This involves converting the code within
the bases of the mRNA into amino acid residues and
each of the 20 possible amino acids is coded for by
three bases in what is called the genetic code. Both
tRNA and rRNA are involved in this process.
GENES AND THE GENOME

The entire complement of double-stranded DNA within a cell
is called the genome, but only about 10% of the DNA in
mammals is thought to carry information in the form of
sequences that are expressed. The sequences of bases
within the long DNA molecules are of two different types:

Those which only occur in single or a few copies, which

are the genes
Regions of repeated sequences, which may be repeated
millions of times within the genome.
Some of these repeated sequences code for proteins
which are needed in large amounts, such as histones. But
the function of much of this DNA is unknown, hence the term
junk DNA. Some of this DNA may be old genes which
have undergone mutations and are no longer transcribed.
However, it has also been suggested that this DNA may act
as a reservoir for the evolution of new genes.
Genome size
The size of the genome varies enormously between organ
isms; the well-studied bacterium Escherichia coli has about
5 million base pairs, compared to the 3 trillion base pairs of
the human genome. In humans there are estimated to be
about 30000 genes, each of which contains the information
for a gene product. Most gene products are proteins; the
only other gene products are the RNA molecules that form
rRNA and tRNA.
2/26/2009 6:48:39 PM

Table 2.16 The genetic code (RNA to amino acids)
First position
Second position
Third position
Phe
Phe
Leu
Leu
Ser
Ser
Ser
Ser
Tyr
Tyr
Stop
Stop
Cys
Cys
Stop
Trp
U
C
A
G
Leu
Leu
Leu
Leu
Pro
Pro
Pro
Pro
His
His
Gln
Gln
Arg
Arg
Arg
Arg
U
C
A
G
Ile
Ile
Ile
Met (start)
Thr
Thr
Thr
Thr
Asn
Asn
Lys
Lys
Ser
Ser
Arg
Arg
U
C
A
G
Val
Val
Val
Val (start)
Ala
Ala
Ala
Ala
Asp
Asp
Glu
Glu
Gly
Gly
Gly
Gly
U
C
A
G
The code given here translates mRNA codons into amino acids; the original sequence of DNA would be complementary to the mRNA and would
contain thymine (T) instead of uracil (U).
THE GENETIC CODE

DNA is only made up of a sequence of four different nucleo
tides (A, G, C and T) which are used to code for 20 amino
acids. Obviously this could not be achieved by one-to-one
coding as, with only four different nucleotides, it would only
be possible to specify four different amino acids. If each
amino acid were coded by two nucleotides, there would still
only be 16 (42) different possible combinations. The way in
which DNA codes for different amino acids uses sets of three
nucleotides to code for each amino acid giving 64 (43) pos
sible combinations. Each group of three nucleotides is called
a codon. The genetic code is usually shown as a table
showing the sequence of three nucleotides in the mRNA
codons and the corresponding amino acid (Table 2.16). It can
be seen from the table that not all of the codons represent
an amino acid. There are three codons that cause the protein
chain to terminate, called stop codons, and one codon
(AUG) that codes for methionine but also acts as a start
codon to initiate the protein synthesis. The codon GUG
usually codes for valine, but occasionally also acts as a start
codon.
All 64 possible codons are used, many amino acids being
coded for by more than one possible codon, but this is not
evenly spread. Some amino acids are represented by six
different codons, whereas others only have a single one. The
code is said to be degenerate. The code is virtually the
same in all organisms and this universality is used as an
argument that life on Earth evolved once only. However,
there are minor differences in the genetic code in mitochon
dria, in some protozoans and in single-celled plants. These
minor changes are thought to be later mutations.
Reading frame
In most cases the code has to be read, in threes, starting at
the AUG codon, to one of the stop codons. This is known as
the reading frame, and most mRNAs are read within a single
frame, although in some cases it has been found that by
starting at a different AUG codon the same mRNA can
produce a different protein. Also some unusual RNAs contain
sites where either four or two bases are read and then the
remaining RNA is read with a frame shift.
ch02.indd 52
Information box 2.13 Deletion of a single base

from DNA will frame shift
the protein
If a single nucleotide is removed from a gene then the

resulting protein will be very different. The protein chain will
be normal up to the point of the deletion. After this all the
resulting codons will be read out of phase. This results in
different amino acids being substituted and the frame shift
will continue adding the wrong amino acids until one of the
new codons corresponds to a stop codon. If the deletion
is near the end of the coding sequence, then the protein
may be nearly normal, but if it is close to the beginning
then changes can be catastrophic. Normal haemoglobin is
composed of two polypeptide chains, - and -globin,
folded to hold the haem molecule. -thalassaemia is an
inherited anaemia characterised by a point mutation in the
-globin chain. The resulting frame shift produces a protein
that contains a series of unexpected amino acids, rendering
the chain unstable and the molecule unbalanced.
Precipitation of the globin chains results in red cells that
are vulnerable to haemolysis, producing a severe anaemia
in those individuals who have inherited the abnormality
from both parents.
TRANSCRIPTION
Transcription is started by the binding of the enzyme RNA
polymerase (RP) to the appropriate section of the appro
priate strand of the DNA (Fig. 2.24). RNA polymerases can
start a new RNA strand (initiation) and extend it (elongation).
In mammals, there are three types of RNA polymerase (RPI,
RPII, RPIII), which make pre-rRNA, pre-mRNA and pre-tRNA,
respectively.
As stated previously, the two strands of DNA are anti
parallel and complementary. Starting at the same end of
each strand would yield two different sequences of bases,
so the RP must bind to the correct strand. The strand serving
as a template is called the antisense strand. The other
strand, the coding strand, is not usually transcribed and is
called the sense strand.
The sequence of bases represented by the sequence of
the mRNA is the same as the original coding strand of the
2/26/2009 6:48:39 PM

DNA rewinding
Sense strand
DNA unwinding
5'
3'
3'
5'
3'
5'
Fig.
Antisense strand
mRNA
2.24 Transcription of DNA into RNA.
DNA, except that mRNA contains uracil instead of thymine.

This is because a complementary copy of a complementary
copy produces the original sequence. This can be followed
through step by step as shown in Table 2.17. It used to be
thought that only the antisense strand was used; however, in
very limited sections of DNA, there are proteins encoded on
both strands.
the others because, being at the beginning of the chain, it

remains a triphosphate. The second base of the DNA then
binds the corresponding free nucleotide, which is then linked
to the first by the RP enzyme. As each nucleotide is added
to the chain, a pyrophosphate is lost, leaving only the single
-phosphate in the backbone of the growing chain. Travel
ling along the DNA in the 3 to 5 direction the RP adds each
successive complementary base to the growing RNA chain.
As the RP proceeds, the DNA unwinds, while areas that have
already transcribed will rewind.
Transcription can proceed quite rapidly at about 40
bases/s. At this rate a very large gene can take many hours
to transcribe. The gene which codes for dystrophin, the
protein that is absent in Duchenne muscular dystrophy, is
more than 2 106bp and would take more than 13 hours to
transcribe. However, most gene transcripts are considerably
smaller. The error rate in copying RNA is about 1 error every
104105 nucleotides. This is high compared to the error rate
in DNA replication but, because RNA is a short-lived mol
ecule and is not repeatedly copied, this rate of error is not
significant.
Starting transcription
Termination of transcription
The antisense strand is read from its 3 end to the 5 end,

and the RNA chain grows from the 5 end to the 3 end.
Each gene has a region called the initiation site, where the
RP must bind to start transcription. In many genes, upstream
(2535bp) from the transcriptional start site is a sequence
called the TATA box, which contains the sequence TATA(A
or T)A, that binds transcription factors that initiate transcrip
tion. Other regions contain areas called promoters, which
are sequences that control the rate at which the gene will be
transcribed. They are located both upstream and down
stream from the gene to be transcribed and bind molecules
called trans-acting factors, which can either enhance gene
transcription or silence it. Many slow-acting hormones, such
as the steroid hormones and thyroid hormones, change the
rate of gene expression, causing changes in the rate of tran
scription of specific mRNAs by forming a ligandreceptor
complex that binds to promoter regions of DNA.
After the gene has been transcribed, elongation terminates

and the RP is dislodged from the DNA strand. How this
occurs in eukaryotes is not yet understood, although it may
involve the formation of hairpin-like secondary structures in
the mRNA that destabilise the RP. Termination often occurs
up to 2000bp past the end of the last exon (see below) of
the gene. The excess mRNA will be ignored, because nothing
after the stop codon is translated. The mRNA has a number
of nuclear proteins associated with it, which are necessary
for the transport out of the nucleus.
Table 2.17 Complementary DNA and mRNA

sequences
Sequence
Direction
Codons or amino acids
DNA coding
strand (sense)
5 3
ATG AGA CTA TTC AGC TAA
Complementary
DNA (antisense)
3 5
TAC TCT GAT AAG TCG ATT
mRNA
5 3
AUG AGA CUA UUC AGC UAA
Amino acids
NC
Met Arg Leu Phe Ser (start)
Spaces are shown between the codons for clarity.
Binding of RNA polymerase

The binding of RP produces separation of the two strands of
DNA in the region of the gene. This separation occurs more
easily between T-A pairs than G-C pairs, because T-A pairs
are joined by two hydrogen bonds whereas G-C pairs have
three. The weaker links of the TATA box may help the DNA
to unwind more easily. The first exposed base then binds the
complementary free nucleotide from the cytoplasm. The first
nucleotide of the RNA chain at the 5 end is different from all
ch02.indd 53
Post-transcriptional processing
All eukaryotic mRNAs are modified after they have been tran
scribed and before they are translated into protein in the
cytosol. While the rest of the gene is being transcribed, and
after only about 35 nucleotides have been added, the primary
transcript of mRNA is capped at the 5 end by the addition
of 7-methylguanosine to the 5 triphosphate group via an
unusual 5-5 linkage. The two 5 end nucleotides may also be
2-methylated. This capping is thought to have a number of
functions that aid further processing and protect the mRNA
from destruction by nuclease enzymes.
Poly-A tails
Next, the 3 end is cleaved at a region called the poly-A site
at the sequence AAUAAA (or rarely AUUAAA). This 3 end of
2/26/2009 6:48:40 PM

the mRNA then has a sequence of up to 300 adenosine (A)
nucleotides added, hence the term poly-A tail. These adeno
sine nucleotides bind proteins to help prevent degradation
of the mRNA. This polyadenylation does not occur in all
RNAs; for example it is absent in histones. However, the
poly-A tails make it easier to isolate mRNAs for experimental
use by fishing for them with molecular probes that consist
of poly-T nucleotides.
RNA splicing: introns and exons

Most eukaryotic genes are much longer than would be pre
dicted by the number of amino acids in the resulting protein.
This is because these genes contain regions, called exons
(short for expressed sequences), which code for the protein,
interspersed with non-coding regions, called introns. Many
genes have multiple introns, which are removed from the
initial primary transcript of mRNA to produce the mature
mRNA.
The final modification to the pre-mRNA involves the
removal of the introns by a process called splicing. This is
done by a proteinRNA complex called a spliceosome. The
positions of the sections to be removed are indicated by
special sequences in the introns which allow the correct
positioning of the different proteins involved in removing the
intron.
Splicing of the same pre-mRNA does not always yield the
same mature mRNA. Exons may also be removed. This alternative splicing can produce mRNAs with different exons,
which will give different proteins. This can be controlled in a
tissue-specific manner. For example, the extracellular protein
fibronectin, which is secreted by different cell types, has two
exons that code for parts of the protein which interact
strongly with cell surface receptors. These are included in
the mRNA produced in fibroblasts but are removed in the
mRNA from hepatocytes. This change results in a functional
difference such that that the hepatocyte fibronectin does
not attach to cells very readily and circulates easily in the
plasma.
RNA editing
This is a recently discovered process, which changes the
sequence of an mRNA. This process is widespread among
protozoans and plants, particularly in the chloroplast. In
mammals, a few examples have been shown. For example,
in a particular protein, by changing a single base in a specific
position from A to G this changes the codon from CAG to
CGG. This changes the amino acid inserted at that position
from glutamine to arginine, with a subsequent change in the
behaviour of the protein.
Processing rRNA and tRNA

The other gene products, rRNA and tRNA, are also processed
before they leave the nucleus. In their mature form, tRNA
molecules contain a number of modified bases. These modi
fications are important in determining the secondary structure
of the tRNAs and the way in which they recognise the en
zymes involved in protein synthesis. Ribose sugars are also
modified by methylation, which makes them more stable.
Assembly of ribosomes
Ribosomal subunits are assembled in the nucleolus, where
the rRNA is produced. Proteins are imported from the cyto
plasm to be combined with the rRNA to make the small and
large ribosomal subunits. The pre-rRNA is split into two
ch02.indd 54
subunits from a single original transcript. This is a way of

ensuring that the separate subunits are produced, as needed,
in equal amounts.
TRANSLATION
After the mRNA has been transcribed and modified in the
nucleus it is transported to the cytoplasm where the pro
duction of the proteins occurs by translation of the mRNA
transcript.
Transport from the nucleus

It has been estimated that more than one million molecules
pass in or out of the nucleus every minute through approxi
mately 4000 pores. These include the mRNAs, as well as
other molecules exported from the nucleus such as ribos
omal subunits and tRNAs. Moving in the opposite direction
are nuclear and ribosomal proteins.
Nuclear pores
After it has been processed in the nucleus, the mature mRNA
is transported to the cytoplasm through one of the nuclear
pores. The normal diameter of the pores is 9nm, but they
can open to a maximum of 25nm. It is thought that mRNAs
that are ready for export include a sequence designating
them for transport to the cytoplasm. Additionally, nuclear
proteins are associated with their 5 end, and these possibly
act as a signal sequence to guide the RNA. The 5 end leaves
the nucleus first, then the nuclear proteins are removed and
returned to the nucleus for reuse.
tRNAs
The function of tRNA is to link a particular amino acid onto
the acceptor stem (see Fig. 2.20) and to bind to mRNA,
through base pairing of three complementary base pairs, the
anticodon, which corresponds to the linked amino acid on
the opposite loop, called the anticodon loop. Because of the
way that tRNA is folded into a clover leaf, the acceptor stem
contains both the 3 and 5 ends of the molecule. The amino
acid is attached to the 5 end of the tRNA in a sequence of
reactions involving the enzyme, aminoacyl-tRNA synthetase.
Using ATP, the enzyme forms an AMP-amino acid, which is
then transferred to the tRNA, releasing AMP. This reaction is
referred to as charging, because the energy from the ATP
bond is passed to the tRNA bond, and is later used to form
the peptide bond, when the amino acid is incorporated into
the protein chain. There are different synthetase enzymes for
each of the amino acids; this ensures that each specific
amino acid is charged with the correct tRNA.
Codon wobble
While there are 61 different codons that code for amino acids
remember that three of the 64 codons are stop codons
there are only about 50 different types of tRNA found in
animal cells. This is also more than the number that would
be required if only one tRNA was used for each amino acid.
The reason for this anomaly lies in the fact that each tRNA
is able to recognise more than one codon, a phenomenon
called wobble (Table 2.18). This happens because the first
base in the anticodon can recognise different bases in the
third position on the mRNA. These non-standard base
pairings are particularly common between G-U bases, which
2/26/2009 6:48:40 PM

Ribosome
Table 2.18 Codon/anticodon base pairings

Third base of the codon
First base of the anticodon
Normal base pairing

A
G
U
C
U
C
A (rare)
G
Unusual base pairing

G
U
A or C or U
U
G
I
I, inosine.
5'
P site
3'
A site
Charged tRNA
Growing
polypeptide
Fig. 2.25 Translation of mRNA.

Protein chains
are formed almost as easily as the normal G-C pairing. Thus

the mRNA codons for phenylalanine, UUU and UUC, are
both recognised by the tRNA anticodon GAA.
Ribosomes and protein synthesis

Ribosomes direct protein synthesis in the cytoplasm in a
cycle that consists of three phases:
1. The initiation phase, when the ribosomal subunits
combine with the mRNA and move to the start codon.
2. The elongation phase, when the mRNA is translated
into protein.
3. The termination phase, when the protein is released
and the ribosome dissociates.
Initiation phase
The first part of the initiation phase is when the small ribo
somal subunit binds to the cap at the 5 end of the mRNA.
It then travels along the mRNA until it reaches the start
codon, AUG. This then binds Met-tRNA and the large ribo
somal subunit then binds to form the complete ribosome.
There are two tRNA binding sites on the ribosome, A and
P (Fig. 2.25). The A site receives the tRNA carrying the next
amino acid to be added to the chain. The P site holds the
tRNA attached to the growing peptide chain.
Ribosomes
5'
3' mRNA
Small
subunit
Fig. 2.26 Polysomes produce multiple copies of a

protein from a single mRNA.
Polysomes
A given mRNA can be translated simultaneously by more
than one ribosome. Once the first ribosome has moved far
enough down the mRNA, the initiation site can be occupied
by another ribosome. This gives rise to a structure called a
polysome, which consists of a single mRNA molecule with
many ribosomes attached to it, each with a lengthening
peptide chain attached (Fig. 2.26).
Lifetime of mRNAs
Elongation starts with the Met-tRNA bound to the P site and

the ribosome assembled. The next codon of the mRNA is in
the A site of the ribosome and is then base paired with the
appropriate tRNA. A peptide bond is formed between the
carboxyl group of the first amino acid, which becomes
the N-terminal amino acid, and the amino group of the
second. The ribosome moves one codon down the mRNA
and the first tRNA is released to be recharged in the cyto
plasm. The second tRNA is shifted to the P site of the ribo
some and a new tRNA occupies the A site. The energy
required to form the peptide bond comes from the tRNAamino acid bond, but the movement of the ribosome and the
movement of the tRNA from the A site to the P site requires
the hydrolysis of GTP to GDP.
A given mRNA can be translated many times until it is

degraded. Enzymes in the cytoplasm gradually remove the
A residues from the poly-A tail. When this becomes too short
then the 5 cap is also removed. The mRNA is then degraded
by exonucleases, which remove nucleotides from each end
of the mRNA. Most mRNAs last a few hours, but some only
last a few minutes and others last a few days. Many of the
short-lived mRNAs code for proteins that are involved in the
control of gene transcription. The half-life of these mRNAs
the time required for the concentration to halve is deter
mined partly by sequences in their genetic code, which allow
them to be targeted for degradation, and partly by regulation
depending on cellular requirements. An example of this is the
mRNA coding for the milk protein, casein. In the presence of
the hormone prolactin, which stimulates milk production, the
rate of transcription of the casein gene rises about threefold.
However, the amount of casein mRNA is increased 100-fold.
This is due to a large increase in the stability of the mRNA,
which in some cases is due to the binding of specific proteins
to sequences in the 3 region after the stop codon.
Termination phase
Rate of protein synthesis
The process of elongation continues, with each step adding

another amino acid to the peptide. When the A site is occu
pied by a stop codon the elongation ceases. The last step,
which also requires GTP, removes the peptide chain from its
attachment to the last tRNA and dissociates the ribosome
into its subunits.
Protein synthesis is a major activity of cells; it is estimated

that, in an average mammalian cell, about one million amino
acids are being added to growing polypeptide chains each
second. A single polypeptide grows at the rate of three to
five amino acids per second. This implies that at any given
time a cell is busy making some 200000 proteins. Clearly,
Elongation phase
ch02.indd 55
Large
subunit
2/26/2009 6:48:40 PM
Information box 2.14 Some viruses break

the rules
In some viruses, the rule observed in cellular organisms,

DNA makes RNA makes protein is not followed. Viruses
can have either DNA or RNA as their genetic material and
they rely on the synthetic machinery of the host cell to
make more copies. In many of the RNA viruses, DNA is not
generally involved in that process as the virus directs the
manufacture of more RNA particles, which also make the
proteins necessary to make more viruses. There are,
however, a group of RNA viruses, called retroviruses, which
use a specific viral enzyme, reverse transcriptase, which is
packaged in the virus along with the RNA, to make a DNA
copy of the viral RNA. This DNA is then incorporated into
the host cell genome, from where it can make more copies
of the viral RNA and proteins.
The most well-known retrovirus is the human
immunodeficiency virus (HIV). One of the consequences of
this packaging of DNA is that the virus can hide in the
genome and remain undetected for years until activated
and new virus particles are produced.
Knowledge of reverse transcriptase enzymes has
revolutionised the study of molecular biology, enabling
researchers to reverse engineer DNA from mRNAs, which
are more easily accessible, allowing the identification of
products.
See also Chapter 5.
Protein production in the cytosol

Proteins for use in the nucleus are transported through the
nuclear pores. These pores are large enough for the protein
to be transported without being unfolded. Nuclear localisa
tion sequences, often rich in positively charged lysine and
arginine residues, interact with the nuclear pores to facilitate
uptake.
Those mitochondrial proteins not made in mitochondria
are manufactured on cytosolic ribosomes. The proteins are
targeted by N-terminal signal sequences, which not only
determine that the protein will be transported into the
mitochondria but also specify whether it will end up in the
outer or inner membrane, or in the matrix or intermembrane
space. Mitochondrial proteins have to be transported through
the membrane unfolded. They are unfolded by molecular
chaperones, transported through the membrane/s, and then
refolded using other chaperones inside the mitochondria.
The enzymes within peroxisomes are made in the cytosol
and imported into the organelles. They are thought to bind
to receptors on the surface of the peroxisomes before
entering through a transport protein. Rare disorders such
as Zellwegers syndrome, which is characterised by central
nervous system, liver and renal abnormalities, seems to
involve a defect in protein import into peroxisomes.
Protein production in the ER

small proteins of 100 to 200 amino acids can be made in less
than a minute. However, many proteins are larger. The largest
protein known, titin, which is found in muscles, has about
30000 amino acids and, at the rate of five amino acids per
second, it would take over 1.5 hours to synthesise each
molecule.
PROCESSING PATHWAYS FOR PROTEINS

As described previously, almost all proteins are modified
after they are translated. Many modifications are not perma
nent but serve to help the targeting of proteins to the appro
priate destinations. As they are produced, proteins are folded
into their final shapes. This is partly determined by their
primary structure and partly by the presence of molecular
chaperones, which aid folding.
Protein can be produced in three different sites:
1. Some proteins are synthesised entirely on free
ribosomes within the cytosol. These are the proteins
that are either destined to remain in the cytosol or
those which are used in the nucleus, mitochondria or
peroxisomes.
2. Proteins destined for the other organelles (ER, Golgi,
lysosomes), insertion into membranes (organelle or
plasma), or for secretion, are synthesised on ribosomes
on the rough ER.
3. Some mitochondrial proteins are synthesised on
ribosomes within mitochondria themselves using
mitochondrial DNA.
Regardless of their final destination, all proteins start
their synthesis on cytoplasmic ribosomes. Their subsequent
fate is determined by the presence (or not) of a signal
sequence of 1630 amino acids at the N-terminal end of
the protein.
ch02.indd 56
For proteins destined for insertion into the rough ER, as the
signal sequence emerges from the free ribosome it is bound
to a complex of protein and RNA, called a signal recognition
particle (SRP). This inhibits further elongation of the protein
until the ribosome reaches the ER. The cytoplasmic surface
of the ER has receptors that bind the SRP and stimulate the
formation of a transmembrane channel, through which the
signal sequence can pass. The SRP then dissociates and is
free to bind another signal sequence. The peptide continues
to grow, but with the N-terminal end being extruded into the
lumen of the ER. The signal sequence is then cleaved by an
enzyme in the ER and degraded. As the protein grows, it
is modified by the addition of carbohydrate and other
residues, folded using chaperones, and when the C-terminal
is reached it is released into the lumen of the ER. The ribo
some is released from the rough ER and dissociates, as does
the transmembrane channel. Many of the varying signal
sequences contain a series of hydrophobic amino acids,
particularly leucine, which help the signal sequence to cross
the membrane. When these are cleaved, the resulting protein
is more hydrophilic and will be retained within the ER.
Transmembrane proteins
Proteins that are destined to be part of a membrane are
inserted into the membrane as it is being made. For a simple
protein with only one transmembrane segment (one end of
the protein inside the cell and one end outside, with the
middle section spanning the membrane), this is achieved due
to the presence of a second signal sequence. The first part
of the protein, the N-terminal end, is made in the same way
as secreted proteins. This will eventually become the external
part of the protein, and it has the signal sequence removed
and is glycosylated as normal. However, when this part
reaches its appropriate size, there is a second signal
sequence that consists of about 22 hydrophobic amino
2/26/2009 6:48:40 PM
Membrane structure and functions 57
Information box 2.15 One type of cystic fibrosis

is due to a protein not
being processed correctly
The most common form of cystic fibrosis is caused by a

deletion of a single amino acid (a phenylalanine at position
508) in the gene that codes for the membrane protein,
CFTR (cystic fibrosis transmembrane regulator). This results
in the ER being unable to export the CFTR protein which is
needed to produce the proper ion channels needed for
cAMP-regulated chloride transport across the plasma
membrane. At normal body temperature, there is an error in
folding of the protein which prevents its export from the
ER. However, curiously, it was found that at lower
temperatures (25C) the proteins fold normally.
These channels are found particularly in the specialised
linings of the bowel and lung. Defective ion transport
results in salt imbalance, drawing water from the airways
and producing the thick lung secretions typical of this
disease. Defective ion transport explains the high
concentration of salt ions in sweat secretions in patients
with CF (the sweat test).
See also Chapter 14.
acids. This forms an -helix, which spans the membrane

and is anchored in it. This signal is called the stop-transfer
membrane-anchor sequence. The transmembrane channel is
disassembled leaving the ribosome to finish the C-terminal
end of the protein on the cytosolic side of the ER.
Protein processing in the ER

In the ER, a number of important steps take place:

Multimeric proteins are formed into oligomers.

Mutated proteins which misfold are identified and
retained.
Unwanted excess protein subunits are degraded.
Asparagine residues undergo N-linked glycosylation.
Precursors of the lysosomal enzymes, prohydrolases,
are glycosylated.

Processing in the Golgi

Proteins move from the ER to the Golgi (Fig. 2.27) where they
are processed further. In the Golgi, as in the ER, there is a
wide range of modifications which occur before proteins can
be sent to their final destinations.
Rough ER
with
ribosomes
Cis Golgi
Medial Golgi
Trans Golgi
Regulated
secretion
Lysosomes
Continuous
secretion
Plasma
membrane
Extracellular fluid
Fig. 2.27 Destinations of proteins.

membrane coated with clathrin bud off from the Golgi, after
which the clathrin is removed. After a number of complicated
steps, involving other vesicles called late endosomes, a
vesicle containing the dephosphorylated enzyme fuses with
the lysosome, and another vesicle containing the empty
mannose receptors returns to the Golgi.
Secretory proteins
Proteins in the Golgi that are destined for secretion do not
have mannose-6-phosphate and are not targeted to lyso
somes. However, they are separated according to whether
or not they are continuously secreted, or their secretion is
regulated (Fig. 2.27). Regulated secretion requires that vesi
cles are stored in the cytoplasm and attached to the cytoskel
eton before they are secreted. A large number of proteins
have been identified that carry out the necessary docking,
priming and fusion. Research carried out in the secretion of
neurotransmitters has helped determine these mechanisms
(see Ch. 8).
Glycosylation
Many of the N-linked sugars that are added in the ER are
modified, and other sugars are added by O-linked glycos
ylation to serine and threonine residues, and then further
modified. Many proteins, especially peptide hormones, are
converted from pro-hormones to their active forms by cleav
age of peptides by proteolytic enzymes. Sphingolipids are
manufactured in the Golgi and glycosylated.
Phosphorylation
The phosphorylation of glycosylated lysosomal prohydrolase
enzymes
generates
terminal
mannose6-phosphate groups, which are then used to target them
to lysosomes. These mannose-6-phosphate groups bind
to receptors on the internal face of the trans-Golgi. These
are clustered into an area of membrane that is coated on
the cytoplasmic side by the protein, clathrin. The areas of
ch02.indd 57
MEMBRANE STRUCTURE
AND FUNCTIONS
FLUID MOSAIC MODEL OF MEMBRANES
A typical cell membrane has hundreds of different types of
protein, which may be pictured as floating in the phospho
lipid sea. This model of proteins embedded in a lipid bilayer,
some of them free to diffuse and some fixed, was originally
proposed by Singer and Nicholson in 1972 and is called the
fluid mosaic model (Fig. 2.28).
It is largely the proteins in a cell membrane which deter
mine its biological function. So, for example, a given cell
responds to a particular chemical signal because its plasma
membrane contains a specific protein. Any cell lacking the
particular protein in its membrane would be completely
2/26/2009 6:48:41 PM

particularly in the ER and in erythrocytes. These can move
phospholipids from one side of the bilayer to the other.
Cholesterol in membranes
Lipid
bilayer
Receptor
protein
Membrane
associated protein
Ion
channel
Fig. 2.28 Fluid mosaic model of membrane structure,
showing integral and membrane-associated proteins.
At normal human body temperature the presence of choles

terol reduces the fluidity of the membrane. It inserts itself
between the phospholipids with its hydrophilic hydroxyl
group facing outwards where the planar structure of choles
terol reduces the movement of phospholipids. The plasma
membrane of cells has more cholesterol than the intracellular
membranes, which makes it more rigid, less permeable to
small molecules and thicker than intracellular membranes.
Some intracellular membranes such as the ER contain very
little cholesterol.
Membrane proteins
The second major component of membranes is protein.
Membrane proteins are of two main types:
unresponsive to the signal. The functions of cell membrane

proteins include:

Enzymes
Transporters
Transmembrane channels
Receptors
Cell markers
Linking the cytoskeleton and the ECM.
Lipid bilayer
All cell membranes, including the plasma membrane, consist
in part of a bilayer of phospholipids, sphingolipids and
cholesterol. Phospholipids are all amphipathic molecules
because they have two acyl groups that are hydrophobic,
and a substituted phosphate, which is hydrophilic because
it is polar. Similarly, sphingolipids and cholesterol both have
hydrophobic and hydrophilic regions. Because of this prop
erty, in an aqueous environment phospholipids aggregate so
that the acyl tails are as far away from the water as possible.
One configuration that permits this is a bilayer.
Movement of lipids in the bilayer

Of itself, a phospholipid bilayer is remarkably dynamic. Indi
vidual phospholipid molecules are free to rotate rapidly
around their long axis and migrate rapidly within their side
of the bilayer. However, transfer of a phospholipid between
the sides of the bilayer (flipping) is very rare. Overall, a phos
pholipid bilayer has the fluidity of olive oil, and it is a highly
impenetrable barrier to most materials. It will, however, allow
the passage of small molecules, such as water, oxygen and
carbon dioxide, and anything that is fat soluble, but it will
exclude almost all polar molecules or charged solutes.
Differential distribution of lipids in the bilayer

Different membranes have different lipid compositions,
including the two leaflets of the same bilayer. An example of
this is the red cell plasma membrane, which has sphingomy
elin and phosphatidylcholine as the major components of the
outer face of the membrane (called the exoplasmic leaflet),
and phosphatidylethanolamine and phosphatidylserine as
the main lipids in the inner side (the cytoplasmic leaflet).
The difficulty with which lipids flip between the two leaflets
helps to maintain this differential distribution. Phospholipids
can be moved between organelles by exchange proteins.
Proteins called flippases are found in some membranes,
ch02.indd 58
Integral (intrinsic)
Peripheral (extrinsic).
Integral membrane proteins can be removed only by

disrupting the membrane structure. Many of these proteins
span the bilayer and have hydrophobic amino acid residues
on the part of the protein in contact with the membrane, while
the parts inside and outside the cell are hydrophilic and may
also carry charged groups. These charged groups are often
glycosylations in the extracellular face, and phosphate
groups on the intracellular face.
Peripheral membrane proteins, in contrast, may be
removed without disrupting the membrane. They may be
associated with the cytoskeleton, or involved in signal trans
duction across the membrane, or they may be attached to
the ECM.
Protein composition of different membranes

The amount of protein varies enormously between different
membranes. The erythrocyte plasma membrane is about half
lipid and half protein with about 8% carbohydrate. In con
trast, the myelin membrane, which surrounds nerve cell
axons and acts as an insulator, is almost 80% lipid. The inner
mitochondrial membrane, which contains all the enzymes
associated with metabolism, is over 75% protein, with no
carbohydrate.
SPECIALISED PLASMA
MEMBRANE STRUCTURES
Many cells, particularly those that make up the solid tissues
of the body, are polarised. That is, different functions are
observed in different parts of the cell. Given the fluidity of the
plasma membrane, and the ability of membrane lipids and
proteins to move laterally within the membrane, in order for
cells to have polarity there needs to be a way of separating
the different areas of membrane. Cells adhere to one another
through interactions of cell adhesion molecules (see below),
but cells that form sheets, separating different body com
partments, must have structures which link them together
with sufficient strength to maintain the integrity of the layer.
Sheets of epithelial cells

Epithelial cells form layers covering most body surfaces.
A typical polarised epithelial cell has an apical surface in
2/26/2009 6:48:41 PM

contact with the lumen (e.g. the intestinal lumen). This may
have one of a number of membrane specialisations (Table
2.19). The basolateral surface is in contact with the basement
membrane, which consists of the basal lamina and the reticu
lar lamina. The proteins in the plasma membrane are restricted
to the apical and basolateral domains by junctions between
the cells (Fig. 2.29).
Junctions between cells

There are a number of different types of junctions
between cells, and between cells and the ECM (Fig. 2.29).
Many sheets of cells sit on a fibrous network of collagens,
proteoglycans, laminin and fibronectin called the basal
lamina.
These junctions can be classified as:
Tight junctions ones that prevent movement of
substances
Adhering junctions ones that maintain cellular
positions
Gap junctions ones that allow movement of
substances.

Transcellular
pathway
Apical
Tight junction
Paracellular
pathway
Microvilli
Lumen
Basement
membrane
Desmosome
Gap junctions
Basolateral
Fig. 2.29 A sheet of epithelial cells.
Table 2.19 Specialisations of the apical membrane

Specialisation
Function
Example
Cilia
Movement across
surface
Bronchial epithelium
Microvilli
Increased area for

absorption
Intestinal epithelium
Stereocilium
Sperm maturation
Epididymal epithelium
Tight junctions
Tight junctions, also known as occluding junctions, form
a belt around the cell attached to the neighbouring cells
known as the zonula occludens. The zonula occludens is
formed from a complex of proteins (ZO1-3, AF-6 and occlu
din) that connect the two membranes tightly together in a
series of studs appearing as interconnected ridges with no
visible extracellular space. Tight junctions are not connected
to the cytoskeleton and prevent the movement of membrane
components between the separated regions of the cell. This
mechanism allows each part to have its own set of proteins
and lipids. Tight junctions also prevent the passage of mol
ecules across the sheet of cells through the route passing
between the cells, the paracellular pathway. This barrier is
produced by a family of proteins called claudins.
Adhering junctions
Three types of adhering junctions, also known as anchoring junctions, are all different types of desmosomes. These
are formed from plaques just below the plasma membrane,
bound to cytoskeletal elements. The plaque is attached to
the plaque of another cell, or to the ECM, by proteins which
span the intercellular space (Table 2.20). Zonula adherens
form a belt around the cell, whereas the macula adherens
only contact the other cell at spots around the cell.
Gap junctions
Gap junctions, also known as communicating junctions,
are formed from integral membrane proteins called con
nexins. A pore in the membrane, called a connexon, is
formed from six connexins, and this is aligned with a con
nexon in the opposing cell. This forms a channel between
the cells which allows the free passage between the cyto
plasms of the cells of water, small molecules (up to 1.2nm
diameter), including some signalling molecules, and ions. It
is this electrical coupling of cells via gap junctions that allows
the movement of excitation across sheets of muscle. This is
particularly important in smooth muscle and cardiac cells.
The connexons tend to cluster in patches on the membrane.
The pore is closed by high concentrations of calcium, which
may be important during apoptosis.
Microvilli and stereocilia

In cells that are required to absorb substances from the
extracellular medium, such as those found in the intestine
and kidney, the surface area for absorption can be increased
up to 30-fold by the presence of microvilli. There may be up
to 3000 microvilli per cell, forming what is known as the
brush border. Microvilli are small (<1m) projections of the
plasma membrane, with a core formed of actin microfila
ments. They are attached to the terminal web, a network of
actin filaments running under the apical surface of the cell.
Stereocilia are longer and branched, with a similar actin core
Table 2.20 Different types of adhering junctions

Adhering junction
Alternative name
Interaction
Linker
Belt desmosome
Zonula adherens
Actinactin
Cadherins
Spot desmosome
Macula adherens
IFsIFs
Cadherins
Hemidesmosome
IFsbasal lamina
Integrin and laminin
IF, intermediate filament.
ch02.indd 59
2/26/2009 6:48:42 PM

to microvilli. They are rare, mainly being found in the
epididymis.
CELLULAR TRANSPORT PROCESSES
Lipid
bilayer
In order for chemical reactions to occur, the reagents must

be placed appropriately along with their enzyme catalysts.
The products and reaction waste products may need to be
moved elsewhere. All cells have a plasma membrane, which
is a barrier to the movement of substances that are not
membrane soluble. The movement of compounds across
this and other intracellular membranes depends on the
chemistry of the molecule and often requires specialised
transport mechanisms.
Low concentration
B
Rate of
transport (J)
Passive diffusion
Small molecules such as water, oxygen, carbon dioxide, and
fat-soluble molecules are able to move through cell mem
branes readily by simple or passive diffusion. Net diffusion
occurs from high concentration to low concentration, that is,
down the concentration gradient. The shorter the distance a
molecule has to diffuse, the quicker the journey will be.
Electron microscopy shows that cell membranes are only
some 9nm across, so the diffusion distance is extremely
short.
This is important for respiration in the lungs, in which
oxygen and carbon dioxide must diffuse through five cell
membranes. To enter the red blood cell, oxygen in the lung
must cross the alveolar cell lining the lungs (two cell mem
branes), the endothelial cell lining the capillary through which
blood flows (two cell membranes) and must then diffuse
across the red cell membrane; carbon dioxide takes the
reverse journey. The total diffusion distance is about 1m
and only 3s is required for this gas exchange to be
completed.
Ficks first law of diffusion

Passive diffusion occurs through a series of random steps,
which result in the molecule being equilibrated between the
compartments. Molecules can move randomly in all direc
tions but the net movement occurs down the concentration
gradient, and the relationship between the rate of transport
and the concentration gradient is linear (Fig. 2.30). Diffusion
across a plasma membrane can be described by Ficks first
law of diffusion:
J = - DA Dc Dx
where J is the net rate of diffusion; D is the diffusion coef
ficient; A is the membrane area; c is the concentration dif
ference across the membrane and x is the thickness of the
membrane.
The diffusion coefficient (D) is defined as:
D = kT ( 6p r h )
where k is Boltzmanns constant; T is the absolute tempera
ture; r is the molecular radius of the diffusing compound and
is the viscosity of the medium.
Diffusion coefficient
From the equation above it can be seen that the diffusion
coefficient (D) is dependent on the viscosity of the diffusing
medium and the molecular radius of the diffusing compound.
The larger the compound and the thicker the medium, the
ch02.indd 60
High concentration
Concentration difference (c)
Fig. 2.30 Passive diffusion: (A) transport and

(B) relationship between rate and concentration difference.
slower the rate of diffusion. An average small water-soluble

molecule, with a diffusion coefficient of 1 105cm/s, will
only take 0.5ms to diffuse 1m (the thickness of a capillary
wall), but it will take 5s to move 100m (the length of many
cells), and 14 hours to diffuse 1cm.
In prokaryotic cells there are no membrane-bound struc
tures within the cytoplasm, so all compounds can diffuse
within the cell to their target. However, over large distances,
this type of transport is very slow, which is possibly one of
the size-limiting factors for both prokaryotic organisms and
within single cells.
Carrier-mediated transport
Many materials that cells need to import or export are either
polar or charged, and so will not diffuse through cell mem
branes. These charged molecules go via special proteins
within the membrane, called transporters. Any transport of
materials that requires transporters is said to be carriermediated transport. Transporters are specific, in the same
way that enzymes are specific; they have preferred sub
strates and the kinetics of transport show the same charac
teristics as enzymes. Transporters are both specific and
saturable, so the maximum rate of transport and the affinity
of the transporter for the substrate can be described using
the MichaelisMenten equation.
There are three types of carrier-mediated transport:

Facilitated diffusion
Active transport
Secondary active transport.

In facilitated diffusion the substrate diffuses from high to
low concentration just as in passive diffusion except that
a transporter is essential for diffusion to occur (Fig. 2.31).
These transporters are protein channels pores, also called
permeases that span the entire width of the membrane. It
is the size and chemistry of the channel lining that allows
specific substrates to pass through it.
2/26/2009 6:48:43 PM

High concentration
A
Other ion channels are opened by small changes in the

electrical potential that exists across the membrane, and
hence are termed voltage-dependent ion channels.
Binding of substrate
Low concentration
Rate of transport (J)
Jmax
Rates of transport
50%
of
Jmax
Km
Concentration
difference (c)
Fig. 2.31 Facilitated transport via an ion channel:

(A) transport and (B) relationship between rate and
concentration difference.
Transport of glucose into red blood cells is one of many

possible examples of facilitated diffusion. Glucose permease
is a transporter found in red blood cells that allows the facili
tated diffusion of glucose. Once inside the cell the glucose
is phosphorylated to glucose-6-phosphate by the enzyme
hexokinase. The transporter does not recognise glucose6-phosphate, and so it remains trapped inside the erythro
cyte. Many of these transporters allow transport in both
directions where the transport is dependent only on the
concentration gradient. Without the action of the hexokinase
glucose would accumulate inside the cell and when the
gradient was reversed could diffuse out through the same
permease.
This permease is one of a number of glucose permeases
called GLUT15 that are found throughout the body. Trans
porters like this, which bind a single substrate, are called
uniporters. The erythrocyte transporter, GLUT1, can trans
port a number of sugars, but each sugar has a different Km.
The Km for d-glucose is about 1.5mM, whereas the Km for
d-galactose is about 30mM. In the presence of both sugars,
the amount of each sugar transported will depend to some
extent on their relative concentrations, but glucose will bind
more readily to the transporter and so will be preferentially
transported.
Ion channels
Another important example of facilitated diffusion involves
the movement of ions. Although charged ions cannot cross
a pure phospholipid bilayer they can diffuse across plasma
membranes under some circumstances via protein pores
called ion channels (see Ch. 4). These channels are selective
for specific ions, so it is possible to talk of sodium channels,
potassium channels and calcium channels. About 40 dif
ferent types of ion channel have been discovered so far.
Many channels are normally closed. Some are opened in
response to chemical signals, either directly (ligand-gated
ion channels) or indirectly (receptor-operated channels).
ch02.indd 61
The mechanism of transport across the membrane using

transporters is thought to involve the binding of the substrate
to a specific site on the extracellular face of the transporter.
This stimulates a conformational change in the transporter
that moves the transported molecule through the protein until
it is exposed on the intracellular face, from which it then dis
sociates. Many of these transporters are able to work in
reverse if the concentration gradient is inverted.
Maximal rates of transport through permeases are in the

order of 102104 molecules per second. Ion channels allow
a faster rate of transfer than do transporters: 107108ions per
second. When the pore is open, more than one ion or water
molecule at a time can enter the channel. This means that
more than one molecule can move through the membrane
simultaneously.
Active transport
Sodium and potassium ions are differentially distributed
across the plasma membrane, with higher concentrations
of sodium outside the cell and higher concentrations of
potassium inside (see Table 2.15). A number of processes,
including diffusion through ion channels, can allow these
concentration gradients to dissipate.
If the only form of transport available was diffusion, both
passive and facilitated, then very quickly all the cellular com
partments would be at equilibrium, with all molecules at the
same concentration throughout the body, and transport
would cease. A second type of carrier-mediated transport
uses the energy contained in ATP to power transport against
the concentration gradient, i.e. to concentrate a compound.
The process is called active transport, and the enzymes that
carry this out are called ATPases, a number of which are
critical for cell function. They use the energy stored in the
high-energy phosphoanhydride bonds of ATP to transport
molecules against their concentration gradients. The ATP is
converted to ADP and the energy released moves the
molecule.
Na+/K+-ATPases
One of the most important of these ATPases is a cation pump
which moves the cations, sodium and potassium, across the
plasma membrane. This is achieved by a transporter called
the Na+/K+-ATPase. This membrane protein takes Na+ from
inside the cell and pumps it out. In exchange, and simultane
ously, the protein pumps K+ from outside the cell into the
cytoplasm. Any transport process that uses metabolic energy
directly, in the manner of the cation pump, is called active
transport, sometimes referred to as direct active transport
because the ATP is used directly by the pump.
The Na+/K+-ATPase consists of four protein subunits,
22, all of which span the membrane and have multiple
transmembrane helices. There are binding sites for ATP on
the internal sides of the subunits, and a site on the outer
surface of the subunits which can bind a number of drugs,
including digoxin (see Information box 2.16). In the most
common type of Na+/K+-ATPase, in each cycle three Na+ are
removed from the cell, and two K+ are added to the internal
2/26/2009 6:48:43 PM

Digoxin
2K+ binding site
3Na+
Na+
Extracellular
Symport
Amino acid
Antiport
Na+
Extracellular
Cell
membrane
Cell
membrane
Intracellular
2K+
3Na+
ADP+Pi
Ca2+ Intracellular
Fig. 2.33 Secondary active transport.
ATP
Fig. 2.32 Na /K -ATPase activity. Pi, phosphate.

+
medium (Fig. 2.32). However, there are Na+/K+-ATPase mol

ecules with other stoichiometries.
The Na+/K+-ATPase cycle is as follows:
1. Pump open to intracellular fluid binds ATP
2. Three Na+ bind to pump
3. ATP phosphorylates subunits
4. Conformational changes and three Na+ released into
extracellular fluid
5. Two K+ bind from extracellular fluid
6. Dephosphorylation of the subunits
7. Pump reverts to previous conformation
8. Two K+ released into cell.
Virtually all cells have cation pumps in their plasma mem
branes. It is estimated that as much as 30% of all ATP gener
ated is used to power cation pumps, a reflection of just how
important they are.
Other ATPases
Another example of active transport is the H+-ATPase found
in the membranes surrounding lysosomes. This transports H+
into the lysosome, reducing the pH to about 5.5, the optimal
pH for the activity of the acid hydrolases. Ca2+-ATPases
are active in removing calcium from the cytoplasm, either
by pumping it into organelles or across the plasma
membrane.
In mitochondria, an H+-ATPase with two subunits F0 and
F1, also known as ATP synthase, runs in reverse, using the
energy of the H+ gradient to produce ATP.
P-glycoprotein
A relatively recently discovered active transport mechanism
explains why some drugs do not cross the bloodbrain
barrier into the brain, despite being theoretically able to cross
the plasma membranes of the capillary endothelial cells. The
transporters, called P-glycoprotein or MDR1 (multidrug
resistance) and MDR2, are present on the endothelial cells
and are able to transport a wide variety of hydrophobic
compounds out of the endothelial cell cytoplasm into the
blood and thereby prevent them entering the brain. This
protects the brain from toxins by preventing hydrophobic
compounds entering the brain by passive diffusion, and any
toxic polar molecules cannot cross the endothelial cells
without a specific transport mechanism.
However, unfortunately, a number of drugs which can be
used to treat cancer, such as vincristine and vinblastine, are
substrates for P-glycoprotein and, because of the activity of
P-glycoprotein, do not enter the brain readily. In many cases,
ch02.indd 62
where tumours become resistant to chemotherapy, this is

because they express a large number of these P-glycopro
tein transporters on their membranes, a process selected for
by continued treatment.
Secondary active transport

A major role of the cation pump is to maintain a large
difference in the concentration of Na+ ions across the cell
membrane. This gradient stores energy in the form of poten
tial energy. A large group of transporters work by allowing
Na+ ions to diffuse from outside to inside the cell, providing
the energy to pump other materials up their concentration
gradients. A good example of this type of transport, called
secondary active transport, is the movement of many
amino acids into cells. Some amino acids are in low concen
tration in the blood, but in higher concentration inside cells.
To import these amino acids into cells requires energy. This
is provided by a co-transport mechanism, which couples
the diffusion of Na+ to the import of amino acids. Many
transport processes harness the energy of Na+ diffusion
in this way, which is also described as indirect active
transport. In this example, transport is active because
energy is required to import the amino acid. Ultimately, the
energy is supplied by ATP, but in indirect active transport
the protein transporter does not, itself, require ATP; the
energy comes from sodium diffusing down its concentration
gradient.
Symport and antiport

There are two types of secondary active transporter (Fig.
2.33). In the example above, the amino acid and the sodium
move in the same direction; this is called symport. In antiport, the sodium moves one way across the membrane
and the other solute moves up its concentration gradient
in the other direction. An example of this is the Na+/Ca2+
antiport found in heart muscle (see Information box 2.16
and Ch. 4).
Coordinated action of transporters

The movement of substances through layers of cells usually
requires the coordinated activity of a number of transport
systems working together (Table 2.21). A good example
of this is the movement of glucose from the lumen of the
intestine into the blood (Fig. 2.34). This entails crossing both
membranes of the epithelial cells which line the lumen. This
requires the activity of three different types of transport:

Secondary active transport by a sodium-linked glucose

transporter
Facilitated diffusion by a glucose permease
+
+
Active transport by the Na /K -ATPase.
2/26/2009 6:48:44 PM

Table 2.21 Summary of transport processes
Type of transport
Transporter/s
Energy source
Rate (molecules/sec)
Passive diffusion
None
Concentration gradient of substrate
Depends on substrate and medium
Uniporter
Ion channel

102104
107108
Active transport
ATPase
Direct ATP ADP
Up to 103
Secondary (indirect) active

transport
Symporter
Antiporter
Indirect ATP ADP

Indirect ATP ADP
102104
102104
Clinical box 2.12 Oral rehydration therapy relies

on the action of transporters
Intestinal
epithelial cells
Lumen
GLUT2
Glucose
Na
Blood
Na+
Glucose SGLT1
Na/K-ATPase
K+
Low
[glucose]
High
[glucose]
Low
[glucose]
Fig. 2.34 Transport of sugars across the intestinal
When diarrhoea causes loss of water and salts from the

intestine, an important part of the treatment involves the
prevention of dehydration, or if this has occurred, the
rehydration of the patient.
Oral rehydration therapy (ORT) is usually formulated as a
powder, which contains a mixture of glucose, amino acids and
sodium chloride, to be dissolved in water for ingestion. When
ORT reaches the intestine, the sodium/glucose and sodium/
amino transport systems are activated, and sodium, glucose
and amino acids are transported across the epithelium. Chloride
ions are drawn through the tight junctions between the cells by
the positively charged sodium ions. This accumulation of
sodium, chloride, glucose and amino acids increases the
osmotic potential in the extracellular space, which then draws
water through the paracellular pathway between the cells and
hence into blood (see also Ch. 16).
lumen.
Information box 2.16 Two transport processes

are involved in the action
of digoxin on the heart
+
The Na /K -ATPase can be inhibited by a number of

compounds extracted from the foxglove (Digitalis purpurea)
called cardiac glycosides, the most important of which
therapeutically is digoxin. This steroid molecule binds to a
site on the external face of the pump and inhibits it. The
consequence is an increased force of contraction of the
heart, which can be of use in the treatment of heart failure.
The increase in force is due to an increase in the
intracellular concentration of calcium. Usually calcium is
removed from the cardiac muscle by the action of a
secondary active antiporter, exchanging external sodium for
internal calcium, a Na+/Ca2+ exchange pump. If the Na+/K+ATPase is inhibited, by the action of digoxin, then the
internal sodium concentration rises and the exchange is
inhibited due to the reduction in the sodium gradient which
powers the transport. Calcium builds up inside the cell and
inactivates the protein troponin, which normally inhibits the
actinmyosin interaction, thus facilitating cardiac muscle
contraction.
See also Chapter 4.
Glucose permease
Glucose then exits the cell into the extracellular space
through the glucose permease, GLUT2, on the basolateral
membrane. This uniporter moves the glucose by facilitated
diffusion down its concentration gradient. It then diffuses
from the extracellular space between the endothelial cells
into the blood.
Na+/K+-ATPase
The sodium, which would otherwise accumulate in the cell
and inhibit further transport, is removed from the cell by
active transport using the Na+/K+-ATPase. This is situated on
the basolateral membrane and ensures that sodium is also
absorbed by this process and is not lost into the gut lumen.
A number of similar transport systems also exist for the
absorption of amino acids, both in enterocytes and in the
epithelial cells lining the renal tubules.
Endocytosis and exocytosis

Sodium-linked glucose transport
The transport of glucose across the luminal brush border of
the intestinal epithelial cells (enterocytes) is carried out by
secondary active transport via a sodium-dependent cotrans
porter called SGLT1 (Sodium/Glucose Linked Transport).
This is an example of a cotransporter where both the sodium
and the glucose travel in the same direction, a symport.
Sodium moves down its concentration gradient into the cell
and glucose moves from the low concentration in the lumen
to the higher concentration in the cell.
ch02.indd 63
Transport using transporter proteins is usually very specific.

Except in the case of P-glycoprotein, there is a limited range
of substrates that can bind to a given transporter. In some
cases only a single molecular species will be transported.
Very large compounds cannot be moved into the cell by this
method, as the energy required to move them through the
membrane is too large. However, molecules and particles in
the extracellular fluid can be internalised by a process called
endocytosis. This involves the enclosure of these external
substances by the plasma membrane, which is pinched off
inside the cell (Fig. 2.35).
2/26/2009 6:48:44 PM
Endosome
Endocytosis
Budding
Docking
Priming
Fusion
Budding
Exocytosis
Ca2+
Fig. 2.35 Endocytosis and exocytosis.
Endocytosis
There are three major categories of endocytosis:

Phagocytosis
Pinocytosis
Receptor-mediated endocytosis.

Phagocytosis
Phagocytosis (cell eating) involves the uptake of large par
ticles into large vesicles called phagosomes. This is mainly
carried out by phagocytic cells of the immune system
macrophages and neutrophils where it is a part of the
bodys defence against pathogens. Phagocytosis involves
the recognition of the bacteria, or other particles, by the
phagocyte, perhaps through antibodies bound to the invad
ing organism (see Ch. 6). This triggers a rearrangement of the
actin cytoskeleton of the phagocytes so that the plasma
membrane is extended around the particle, forming an intra
cellular membrane-bound organelle called a phagosome.
Once the particle is engulfed, the phagosome is moved using
motor proteins to the lysosomes, where the acid hydrolases
can break down the particle into its component parts. These
can then be utilised by the cell. Anything that cannot be
degraded remains in the lysosome as a residual body. In
long-term smokers, for example, lung tissue is blackened by
the accumulation of macrophages in the septa of the alveoli
that contain black substances derived from inhaled smoke.
Pinocytosis
Pinocytosis (cell drinking) is carried out by small vesicles
(pinocytes) that take up external fluid and solutes. Cells can
also ingest large amounts of bulk fluid and solutes by macro
pinocytosis, where an extension of the plasma membrane
spreads around an area of extracellular fluid that can be
large, up to 100nm. For example, cells in the thyroid gland
use this method to take up thyroglobulin.
Receptor-mediated endocytosis
Many compounds present in the extracellular fluid, especially
those that are too large to be taken up by transporter pro
teins, can be specifically transported by receptor-me diated
endocytosis. This is a mechanism for concentrating and
ch02.indd 64
internalising substrates, often proteins, glycoproteins or car

bohydrates. It is also referred to as clathrin-dependent
endocytosis, as it involves the formation of vesicles in spe
cific areas of cell membrane that are coated with an intracel
lular lattice of a protein called clathrin. The substrate binds
to cell surface receptors, which are clustered on the area of
membrane marked by the clathrin. The binding of the ligand
causes the membrane to fold inward, forming a coated pit.
The edges of the pit fuse together and an intracellular vesicle
coated with clathrin is formed. The clathrin then dissociates
from the vesicle, now called an early endosome. This fuses
with another endosome, which has a low internal pH (about
5), becoming a late endosome, or CURL. The low pH is due
to the active transport of H+ into the enzyme by a H+-ATPase.
Due to the low pH the ligands then dissociate from their
receptors. The membrane, with its inserted membranes, is
returned to the cell surface and the ligand is delivered to a
lysosome, where it is either broken down into the required
components or transported into the cytoplasm.
Two substances taken up by most cells using receptormediated endocytosis are low-density lipoproteins (LDLs),
which carry about 75% of the plasma cholesterol around the
body, and transferrin, which carries iron into the cell.
LDL particles are spheres, about 20nm in diameter, com
posed of a single phospholipid layer plus some cholesterol,
surrounding a core of cholesterol esters (cholesterol plus a
fatty acid). Embedded in the phospholipid monolayer is a
single protein, called apoB, which binds to LDL receptors on
the cell surface. Once bound to LDL receptors, the LDL
particle is ingested in a clathrin-coated vesicle. The LDL
receptor is eventually recycled to the cell surface and the
LDL particle is degraded into cholesterol, fatty acids and
amino acids (from the apoB protein). Cells balance their
requirement for cholesterol by regulating both the synthesis
of cholesterol and the number of LDL receptors in their
surface.
Another example of receptor-mediated endocytosis, fol
lowed by sorting in endosomes, is the transport of transfer
rin. Transferrin receptors bind ferrotransferrin, a combination
of the iron-binding protein, transferrin, found in plasma,
along with bound iron. Ferrotransferrin is endocytosed in
clathrin-coated vesicles. However, after the fusion with a late
endosome, the transferrin is not dissociated from the recep
tor, but iron is dissociated from the transferrin. This allows
the iron-free apotransferrin to be recycled to the plasma
when the transferrin receptors are returned to the plasma
membrane.
Exocytosis
The process by which the receptors are returned, along with
their membrane, is the inverse of endocytosis, a process
called exocytosis. This involves the fusion of intracellular
vesicles with the plasma membrane, releasing their contents
into the extracellular fluid. Exocytosis is not only the mechan
ism by which endocytotic receptors are returned to the cell
surface but it is the main mechanism by which hormones and
neurotransmitters are secreted from cells (see Ch. 8).
Transcytosis
Some molecules are moved across a sheet of cells, such as
the intestinal epithelia, by a combination of endocytosis and
exocytosis, called transcytosis. The molecules are moved
by receptor-mediated endocytosis into endosomes, which
travel across the cell and fuse with the opposite plasma
2/26/2009 6:48:44 PM
Cell-to-cell communication 65
Information box 2.17 High levels of plasma

cholesterol can be
caused by a defect in
LDL receptors
A relatively common genetic disorder known as familial

hypercholesterolaemia, causes a raised level of cholesterol
in the blood. This defect occurs in about 1 in 500 of the
normal population and is associated with a family history of
early heart disease.
There are a number of genetic mutations responsible for
causing the disease, but the common theme is a reduction
in the clearance of LDL particles via the LDL receptors. In
some cases the receptor numbers are reduced. In others it
is the receptor which is defective.
Another form of familial hypercholesterolaemia that is
relatively common is caused by a defect in the apoB
protein which prevents binding of the LDL particles to the
receptor.
While many heterozygotes (who only carry one copy of
the defective gene) have mild symptoms, children who have
inherited the disorder from both parents are severely
affected, most dying from heart disease before adulthood if
untreated. Even in those treated individuals about 50% will
die by the age of 60.
See also Chapter 3.
Paracrine
Target
cell
Local
hormone
Autocrine
Receptor
Hormone or
growth factor
Neural signalling
Stimulus
Release of
neurotransmitter
Electrical signal
Target cell
Neurotransmitter
receptor
Endocrine
Stimulus
membrane, where the molecules are released. The cycle is

completed by the endocytosis of the vesicle membrane
containing the receptors and its exocytosis on the other side,
returning the receptors to their original place. An example of
transcytosis is the movement of immunoglobulins across the
intestine of the newborn from the mothers milk. Interestingly,
the receptors will bind the immunoglobulins at the slightly
acid pH (pH = 6) of the intestinal lumen, but release them
at the neutral pH of the interstitial fluid. These ingested
immunoglobulins confer significant levels of immunity to the
newborn while its own immune system is developing.
Bulk flow in the blood

Diffusion and membrane transport processes are sufficient
for moving substances short distances, but for large dis
tances the time required would be impossible. The fastest
way for substances to move around the human body is in
the blood. Propelled by the contraction of the heart, the bulk
flow of blood, with its suspension of cells, lipid particles and
proteins, allows the movement of large quantities around the
body at high speed. In humans, this bulk flow can move
oxygen from the lungs to the furthest blood vessels in the
limbs in 30s, from where it can diffuse the 10m or so into
the tissues (see Ch. 11).
CELL-TO-CELL COMMUNICATION
The human body is a large structure that needs to behave in
a coordinated fashion. This can be achieved through signal
ling mechanisms, which transfer information about biochem
ical events between cells. Signalling requires two interacting
elements:

ch02.indd 65
A signalling molecule
A receptor molecule so the signal can be recognised
and acted upon.
Hormone
Blood
Receptor
Target
cell
Fig. 2.36 Different types of cell signalling.

The signal molecule, or ligand, binds to the receptor at a
specific binding site, causing a change in the receptor, which
can produce an effect in the target cell, the effector. This
process is called signal transduction.
TYPES OF SIGNALLING
There are four main forms of signalling (Fig. 2.36):
Paracrine: this is where molecules called local
hormones are released into the local environment of the
cell producing the signal. These act on cells in the
immediate environment.
Autocrine: these molecules act on the cell releasing the
signal. This is the mode of action of many growth
hormones. Unregulated release of growth hormones can
lead to the formation of tumours.
Neural signalling: this is a particular type of signalling
that can be both paracrine and autocrine. A signal
generated in a nerve cell travels along nerve fibres to
the effector. These signals travel electrically down these
fibres until the target is reached, where a chemical
called a neurotransmitter is released locally (see ch. 8).
This signal can act either on the releasing nerve cell
(autocrine), or on the nearby target (paracrine), which
may be an effector, such as a muscle or gland, or
another nerve.
Endocrine: the signal is called a hormone and is
released into the blood where it can circulate to the
whole body. However, it will only affect cells and tissues
that have receptors for the hormone (see Ch. 10).

2/26/2009 6:48:45 PM

Another type of local signalling occurs when proteins in
the cell membrane are attached to receptors on adjacent
cells. These are involved in cell-to-cell recognition and cell
adhesion.
While paracrine and autocrine signalling rely on the
passive diffusion of the signalling molecule and so are only
active over a small distance, both neural and endocrine
signalling allow information to be transmitted over large dis
tances in a short time. Hormones released into the bloodstream can travel throughout the body within seconds, and
although neurotransmitters themselves only have to diffuse
very short distances, the electrical signals which trigger their
release can travel for many metres. The longest nerves in the
human body connect the lower spinal cord to the toes; the
same nerves in a giraffe are many metres long.
LIGANDS AND RECEPTORS

The ligands involved in cell signalling are extremely varied.
Some are small molecules derived from amino acids, such
as noradrenaline, which is synthesised from tyrosine, while
others are proteins, like insulin. An unusual ligand only
recently discovered is nitric oxide (NO). Known previously
only as a poisonous gas, this is now known to be released
from endothelial cells (and some neurons) where it acts
locally. It also has a role in the immune response, where it
combines with superoxide anions to produce a compound
poisonous to bacteria and parasites.
Many molecules can act as ligands for more than one
type of signalling (Table 2.22). For example, noradrenaline
acts as both a hormone and a neurotransmitter. It is released
from many nerve terminals in the autonomic nervous system
(see Ch. 4) but it can also be released into the blood from
the adrenal medulla.
Receptors, like enzymes and transporters, show speci
ficity in their binding and the kinetics of binding can be
Clinical box 2.13 Nitric oxide relieves the pain
of angina
In 1867 the pharmacologist Lauder Brunton observed that amyl
nitrite inhalation relieved the pain of angina, but the mechanism
of this effect was unknown. Amyl nitrite has since been
superseded by nitroglycerin (glyceryl trinitrate), which acts by
being degraded in the blood, releasing nitric oxide. This acts to
relax smooth muscle cells found in blood vessel walls, thus
increasing the diameter of the vessel and increasing blood flow.
The pain of angina is due to the heart muscle not receiving
enough oxygen delivered in the blood; increasing the blood
supply through vasodilation thus reduces the pain.
described in a similar way to transport kinetics using the

MichaelisMenten equation. Specific receptors also mediate
a specific response.
Receptors fall into two main categories depending on
their ligands:
Cell surface receptors: these are integral membrane
proteins with an extracellular binding site for the ligand.
The ligands for these receptors are either hydrophilic
molecules, which cannot diffuse across the cell
membrane, or are molecules which are too large to be
easily transported across the membrane.
Intracellular receptors: these bind to their ligands
either in the cytosol or in the nucleus. The cytosolic
receptors then translocate to the nucleus. Many of the
ligands for these receptors are the steroid hormones,
which are derivatives of cholesterol, and can diffuse
across the cell membrane. Others include thyroid
hormone, which is transported into the cell as an
inactive prohormone (T4) and is subsequently converted
into the active form (T3), which binds to nuclear
receptors, and the retinoids which include vitamin A.

Cell surface receptors

The cell surface receptors fall into three main groups

G-protein-coupled receptors
Ligand-gated ion channels
Receptor kinases.

G-protein-coupled receptors
The most abundant type of cell surface receptors are called
either G-protein-coupled receptors (GPCR) or 7TM receptors. Both of these names derive from the fact that they all
share a similar transduction mechanism, changing intracel
lular biochemistry via a group of proteins called G proteins,
which bind guanine nucleotides (GTP and GDP). The recep
tors also share a similar structure, in that they consist of
an integral membrane protein with seven transmembrane
segments, with an extracellular binding site for the ligand and
an intracellular region that interacts with the G protein. They
are also called metabotropic receptors because of their
actions on intracellular metabolism. Many of these receptors
exist in multiple isoforms. For example, the receptors which
respond to smells are GPCRs; in humans there are thought
to be over 500 different genes which code for GPCRs (com
pared to over 1000 in mice).
Table 2.22 Some cell signalling molecules

Type of signalling
Example
Example of effect*
Paracrine
Glucagon
Nitric oxide
Increases insulin release

Local vasodilator released from endothelial cells
Autocrine
Prostaglandins
Noradrenaline
Inflammatory mediator
Regulation of noradrenaline release via autoreceptors
Neural
Glutamate
Acetylcholine
Noradrenaline
Excitation of nerves in the central nervous system

Excitation of skeletal muscle
Vasoconstricts blood vessels
Endocrine
Adrenaline/noradrenaline
Glucagon
Testosterone
Increases heart rate

Increases blood glucose
Male sex hormone
*Many of the compounds listed have multiple effects, only a single example is given here.
ch02.indd 66
2/26/2009 6:48:45 PM
Cell-to-cell communication 67
Table 2.23 Some G proteins and their associated enzymes and second messenger systems
G protein family*
Enzyme affected
Second messenger (or effect) produced
Other effects
Gs
Adenylate cyclase
Increased cAMP
Opens Ca2+ channels and closes Na+

channels
Gi
Adenylate cyclase
Decreased cAMP
Closes Ca2+ channels and opens K+

channels
Gq
Phospholipase C
IP3, DAG
IP3 releases intracellular Ca2+ from stores

which acts as another second messenger
Go
Phospholipase C
IP3, DAG
IP3 releases intracellular Ca2+ from stores,

closes Ca2+ channels
Gt
cGMP phosphodiesterase
cGMP
*Strictly, these are the names of the -subunit family of the G protein.
cAMP, cyclic adenosine monophosphate; IP3, inositol trisphosphate; DAG, diacylglycerol; cGMP, cyclic guanosine monophosphate.
G proteins
The G proteins are activated by the receptors and in turn
activate other intracellular signalling processes, often involv
ing the production of intracellular signalling molecules called
second messengers. Despite the very wide range of differ
ent GPCRs, the number of different second messengers
produced seems to be limited. Some of the best known are
cAMP, cGMP, calcium, inositol trisphosphate (IP3) and diacyl
glycerol (DAG).
The G proteins in their inactive form are trimers consist
ing of , and subunits. When they are activated by the
receptors, the subunit separates from the and subunits
(which remain together). The subunit then interacts with
an effector, which is often an enzyme, but may also be a
membrane ion channel. The effect of the G protein is to a
large extent determined by the identity of the subunit.
There are known to be at least 16 different genes coding for
subunits, which fall into five or six groups (depending on
the system used) according to their actions and locations
(Table 2.23). This means that many different receptors must
activate the same G protein. The different G proteins affect
only a small number of intracellular enzymes. These enzymes
produce the second messenger molecules, which them
selves activate protein kinases and phosphatases to influ
ence cell biochemistry.
Light receptors are GPCRs

As well as being responsible for the sense of smell, GPCRs
also mediate vision where the ligand is a photon of light (see
Ch. 8). In humans there are four types of light-sensitive GPCR
in the eye. One of these, rhodopsin, absorbs a wide range
of photons at low intensity and is responsible for black and
white vision in cells called rods. Colour vision is mediated
by three types of coneopsin, which preferentially absorb
either red, green or blue light. These are found in cone cells,
and in the brain the signals from these three kinds of receptor
are processed in a way that identifies the 16 million different
colours which it has been estimated that humans can
distinguish.
Ligand-gated ion channels

Another major group of cell surface receptors are the ligandgated ion channels. These are multimeric integral mem
brane proteins that have an extracellular binding site for the
ligand. Binding of the ligand causes a conformational change
in the protein which opens up a pore that crosses the mem
brane, through which ions can move. Each type of ligandgated receptor allows a specific ion to move through it, and
ch02.indd 67
this is determined by the amino acid sequence of the amphip

athic helices which line the pore. An example of this type of
receptor is the acetylcholine receptor found on all skeletal
muscle. When this is activated by acetylcholine, a pore
opens, which allows Na+ to flow into the muscle cell and
depolarise it, a process which initiates muscle contraction
(see Ch. 9).
Receptor kinases
Other cell surface receptors activate intracellular tyrosine or
serine/threonine kinases. These kinases may either be an
integral part of the receptor, as in the case of the insulin
receptor, or in the case of the cytokine receptors the receptor
may activate cytosolic tyrosine kinases. Other receptors,
such as the receptor for atrial natriuretic peptide, activate a
different membrane-integral enzyme, guanylate cyclase, to
produce cGMP. Guanylate cyclases are also the target for
nitric oxide, although these ones are cytosolic. Nitric oxide
activates them to increase cGMP levels, which in turn leads
to smooth muscle relaxation. Other receptors for growth
factors and cytokines activate pathways that influence gene
activity via transcription factors.
Intracellular receptors
The intracellular receptors for the steroid and other intra
cellular ligands are very different from the cell-surface
receptors. When activated they all affect gene transcription
because the receptors bind directly to DNA. These recep
tors consist of large monomeric proteins with a hormonebinding site and a region of about 60 residues, called the
DNA-binding domain, which has a similar structure in all
of the different receptors. It contains two loops of about 15
residues, called zinc fingers, which contain four cysteine
residues surrounding a zinc atom. It is these fingers that
are thought to bind to the DNA. Binding of the ligand to
the receptor causes the protein to form dimers, which then
bind to a specific sequence on the DNA, found about 200
base pairs from the start of the appropriate gene, the
hormone-response element. The binding of the receptor
ligand complex causes an increase in transcription of the
gene.
Nuclear and cytoplasmic receptors

Thyroid hormone receptors are found in the nucleus, perma
nently bound to DNA, but the steroid receptors are found,
initially, in the cytoplasm. When they have bound their ligand
they move to the nucleus and bind to DNA.
2/26/2009 6:48:46 PM

CELL RECOGNITION
In multicellular organisms, cells must have a way of interact
ing with the surrounding cells and the extracellular compo
nents. This is particularly important during development,
where cells are forming tissues and there are considerable
movements of cells from place to place. However, even in
the adult, cells need to divide and grow in order to replace
themselves and respond to changes in demands. In order for
cells to grow in a controlled manner they need to be aware
of their surroundings. They do this by means of a small
variety of cell adhesion molecules. Many of the cellcell
interactions are relatively permanent, as in muscle, which
must bind to other muscle cells and to tendons, and in skin,
where cells must adhere to each other and the underlying
matrix. Other interactions are much less permanent, such as
that between capillary endothelial cells and white blood cells
(see below).
CELL ADHESION MOLECULES

Four main types of molecules allow cells to make connections
between each other and the ECM. Firstly, the two types
of calcium-dependent molecules, the cadherins and the
selectins, and secondly, the calcium-independent molecules,
the integrins and the immunoglobulin superfamily of cell
adhesion molecules known as CAMs. They are all transmem
brane proteins with intracellular and extracellular domains.
Homophilic binding occurs when both ligand and recep
tor are the same molecule. Heterophilic binding joins two
different molecules, although which is referred to as the
receptor and which as the ligand is not important. As well as
these major types there are a number of other adhesion
molecules that do not fall into these four groups.
Cadherins
There are more than 80 different types of cadherins so far
identified. The intracellular domain interacts with actin via
proteins called catenins, and the extracellular section con
sists of a protein dimer with four calcium-binding sites. The
dimer then binds to cadherin dimers on the opposing cell.
E-cadherins are found on most epithelial cells, while
N-cadherins and P-cadherins are found on neural (plus
some muscle) and placental cells, respectively. The removal
of calcium reduces cell adhesiveness, a factor taken into
account when preparing tissue extracts for cell culture.
Selectins
Selectins belong to a group of molecules called lectins,
which are proteins that bind to carbohydrates. They bind to
the carbohydrate portion of glycoproteins, especially on the
surface of white blood cells (leucocytes), helping them to
target sites of inflammation. One of the selectins, P-selectin,
is usually found in vesicles inside endothelial cells lining the
blood vessels. When the endothelial cell is stimulated by
factors released during inflammation, these selectins are
inserted into the cell surface membrane, where they can
interact with circulating leucocytes. The leucocytes can then
move across the endothelial cells to reach the required
tissue.
Integrins
The integrins are mainly used in interactions between the
cell and the ECM, binding to adhesion proteins in the ECM
ch02.indd 68
Information box 2.18 HIV infection involves the

virus binding to a specific
cell surface molecule
The cells of the immune system that are targeted by the

HIV virus are called T helper cells. The reason that these
particular cells are infected is the presence on their surface
of a protein of the IgG superfamily called CD4. The virus
attaches to the T cell via a protein on the viral surface
called gp120.
called laminin and fibronectin. These, in turn, bind to the

proteoglycans of the ECM, ensuring that the cells and the
ECM are bound together. Inside the cell the integrins bind
to cytoskeletal proteins. Many tumour cells do not have
fibronectin on their surface, aiding their migration through the
ECM. This is a critical factor in their ability to spread to other
parts of the body (metastasis).
CAMs
The cell adhesion molecules known as CAMs have a similar
structure to the G-type immunoglobulins (IgG). They can bind
homophilically or heterophilically to other CAMs. N-CAMs
are found on neural cells where they mediate adhesion, while
intercellular (I)-CAMs and vascular (V)-CAMs are involved in
the interactions with leucocytes.
CELL DIVISION AND DNA

REPLICATION
Proteins are continually being replaced inside cells and, at
the same time, cells themselves are continually dying and
in many cases being replaced. Cells have many different
forms and mature cells are said to be differentiated.
The replacement of cells is carried out by one of two
processes:
An undifferentiated precursor cell, a stem cell, is
stimulated to develop into the required cell type.
A fully differentiated parent cell divides into two by a
process called mitosis, producing two smaller daughter
cells which then grow into two parent cells.
Another type of cell division, called meiosis, is required

to produce the gametes, e.g. the eggs and sperm of
mammals. Both mitosis and meiosis require DNA
replication.
STEM CELLS
Some cells do not undergo mitosis. Once formed, mature
erythrocytes and nerve cells cannot divide they are post
mitotic. Other cells will undergo mitosis, although the rate at
which this occurs varies enormously. At the other end of the
spectrum, stem cells divide continually and can be stimu
lated to differentiate into mature cells. Some stem cells can
produce all of the different cell types in a tissue they are
pluripotent. Others are multipotent, that is, they can
produce some but not all of the cells in a tissue.
In different adult tissues there are stem cells that can be
induced to form the different cells of which that tissue is
2/26/2009 6:48:46 PM
Cell division and dna replication 69
Information box 2.19 Stem cells may be used to

produce replacement
tissues
While adult stem cells from different tissues may be used to

produce replacement cells of that tissue type, there is a
great deal of interest in the use of embryonic stem cells,
which produce all the different tissues of the embryo
(except the placenta). One line of current research aims to
treat a type of diabetes (juvenile onset type 1), where the
insulin-producing -cells of the pancreas are destroyed, by
replacing them with -cells derived from stem cells. Other
studies are looking at replacing damaged heart muscle
using stem cells. However, the advantage of using adult
stem cells from the patients themselves ensures that there
are no problems with rejection and therefore no need for
immunosuppressive drugs.
1 Prophase
Nuclear envelope
Centromere
Chromatid
2 Metaphase
3 Anaphase
4 Telophase
G1
(Gap 1)
Restriction
point
Fig. 2.38 Phases in mitosis (for clarity only three

G0
most cells, once they pass the restriction point are commit
ted to dividing.
M (Mitosis)
G2 (Gap 2)
(DNA synthesis)
S
Fig. 2.37 The cell cycle.
made. For example, in the bone marrow, pluripotent stem

cells can divide and form the red blood cells, platelets and
the full variety of white blood cells (leucocytes) that are found
in the blood. Bone marrow transplants from donors, which
contain these stem cells, require matching of the major cell
surface antigens in order to avoid donorversushost reac
tions. Some tissues only have very few stem cells, although
there is some evidence that the numbers may increase
following stress.
PHASES OF THE CELL CYCLE

The term cell cycle describes the life cycle of a cell from the
time at which it is produced to when it divides into two
daughter cells. The cycle consists of two major parts: mitosis
and interphase (Fig. 2.37). During mitosis, also called the M
phase, the cell divides rapidly. In a cell which is dividing every
24 hours, the M phase lasts about 1 hour. Interphase is
divided into three phases, G1, S and G2.
In the absence of specific growth factors there is a point
in late G1, called the restriction point, when the cell enters
a quiescent phase called G0. However, if the appropriate
growth factors are produced, the cell continues through G1
until it reaches the S phase where the DNA of the cell is
replicated. This produces a cell containing two copies of the
DNA, one for each of the daughter cells. This process takes
about 8 hours. The cell then remains in G2 until it divides.
There are a very few cell types which can remain in G2, but
ch02.indd 69
chromosomes are shown).
The human karyotype

The number and type of chromosomes in a particular species
or individual is called the karyotype. Humans normally have
46 chromosomes: 22 pairs of autosomal chromosomes,
which are the same in males and females, and a pair of sex
chromosomes. Normal males have one Y chromosome and
one X chromosome, whereas normal females have two X
chromosomes. The cells are said to be diploid because they
have 2n chromosomes, where n is the number of chromo
some pairs. So in humans n is 23.
In mitosis, a single cell that has already duplicated its
DNA, so that it contains twice the normal amount (4n), divides
into two virtually identical daughter cells. Each of the daugh
ter cells is diploid: it has its own nucleus, which contains the
normal amount of DNA (2n), and half the cytoplasm and
cytoplasmic organelles of the parent cell.
In meiosis, the starting point is the same: a cell with 4n
chromosomes. However, this is followed by two cycles of
cell division, so that the resulting cells are haploid (1n).
Sexual reproduction requires the fusion of two gametes, one
from each parent. When female and male gametes (an egg
and a sperm) combine they each contribute 1n chromo
somes to produce a diploid (2n) fertilised ovum or zygote.
MITOSIS
Mitosis is the division of the single nucleus into two identical
daughter nuclei and is divided into four periods (Fig. 2.38):
1. Prophase. This begins when the replicated DNA
condenses into visible chromosomes. This condensation
involves each chromosome becoming more and more
coiled until it is dense enough to be observed under a
light microscope. The nucleoli disappear and the
nuclear envelope breaks down. Each pair of replicated
chromosomes, called sister chromatids, are joined at a
2/26/2009 6:48:46 PM

characteristic point where the chromosome appears to
be constricted, called a centromere. The centrioles,
which are also duplicated during interphase, move apart
in pairs, towards opposite ends, or poles, of the cell to
form a centrosome, or microtubule-organising centre,
with a radiating array of microtubules that is visible
between the two centrosomes forming the mitotic
spindle.
2. Metaphase. In the region of each centromere
develops a kinetochore, which attaches the
chromosome to the microtubules. The chromosomes
align themselves on the microtubules at the centre of
the cell, the equatorial or metaphase plate. These
kinetochore microtubules are used to pull the
chromosomes one from each pair towards
opposite poles of the cell.
3. Anaphase. This occurs when the centromeres of each
chromosome pair split and the kinetochore microtubules
shorten, pulling each chromosome to opposite ends of
the mitotic spindle. This is balanced by other
microtubules (radiating microtubules) that anchor the
centrosome to the plasma membrane. Once the
chromosomes have reached the centrosomes, the
poles separate further by elongation of the polar
microtubules.
4. Telophase. This is marked by the uncoiling of the
chromosomes and the re-formation of a nuclear
envelope around each group of chromosomes. Around
the equator of the cell an indentation forms where a
contractile ring, formed from actin and myosin, pinches
the cell until it cleaves into two daughter cells.
Strictly speaking, mitosis is the division of the nucleus
only. Cytokinesis is the name given to the division of all of
the cell except for the nucleus. Mitochondria, ribosomes and
other organelles need not be distributed equally, just as long
as each cell has some of them, and there is no mechanism
which ensures their equal distribution. Usually the cytoplasm
is divided equally between the two daughter cells, but this is
not always the case.
The replication of mitochondria occurs throughout inter
phase. As mitochondria grow in size, other mitochondria
are pinched off in a similar way to the division of some
bacteria.
Daughter cells may not be completely identical to each
other. This may be because they have different mitochondria,
which do not all have identical DNA but also because DNA
replication may not be perfect and one of the daughter cells
may carry a mutation. Furthermore, if the chromosomes are
not divided correctly between the two daughter cells, then
the resulting cells will have an abnormal number of chromo
somes, which is called aneuploidy.
As only a small proportion of cells in a population are
dividing at any one time, it is difficult to observe the chromo
somes. However, cells can be blocked at metaphase by the
alkaloid colchicine, which prevents the chromosomes divid
ing. Cells then accumulate in metaphase and the chances of
observing the chromosomes are much higher.
MEIOSIS
While mitosis produces virtually identical daughter cells
with a full complement of chromosomes, during meiosis
(Fig. 2.39) two processes occur.
ch02.indd 70
1a Early prophase 1
A
A
1b Late prophase 1
AA
B
2 Metaphase 1
B
A A
A A
or
B B
B B
3 Anaphase 1
4 Telophase 1
5 Prophase 2
6 Metaphase 2
7 Anaphase 2
8 Telophase 2
Fig. 2.39 Phases in meiosis.

Prior to meiosis, during interphase, the DNA is replicated
(just like mitosis), which increases the number of chromo
somes to 4n, four of each type. But, unlike mitosis, this is
followed by two cycles of cell division. These are called
meiosis I and II and each consists of the same four phases
as in mitosis. This produces cells with half the number of
2/26/2009 6:48:47 PM

Chromatids
3'
5'
Homologous
chromosomes
Leading
strand
DNA polymerase III
5'
Primosome
3'
DNA polymerase I
Chiasma
Lagging
strand
3'
Recombinant
chromatids
5'
RNA primer
DNA polymerase III
Okazaki fragment
Fig. 2.41 DNA replication.

Fig. 2.40 Recombination during prophase 1 of
meiosis.
chromosomes (1n), one of each of the 22 autosomal pairs
and either an X or Y chromosome.
During meiosis, genetic material is exchanged between
chromosomes in a process called recombination, or crossing over, so that each resulting gamete has a different com
bination of genes. This is what produces the genetic diversity
between individuals, as each person, while inheriting their
parents genes, gets them in a unique combination.
The phases of meiosis

1. Prophase 1. Firstly, the chromosomes condense, with
each chromosome consisting of two chromatids joined
by a centromere. The homologous pairs of chromatids
form pairs along their whole lengths, a process called
synapsis. During this time, recombination exchanges
parts of chromatids. This can be seen by the formation
of structures called chiasmata between pairs of
chromosomes (Fig. 2.40). This recombination allows
for novel combinations of genes to occur in the
chromosomes. In late prophase 1, the nuclear envelope
breaks down.
2. Metaphase 1. The pairs of chromosomes line up on the
equator of the cell and attach to the microtubules via
the kinetochores.
3. Anaphase 1. The chromosomes separate, with one pair
of chromatids going to each pole.
4. Telophase 1. The chromosomes form into nuclei and
the nuclear envelope starts to reform. The cell then
divides into two.
5. Prophase 2. After a very brief interphase, when the
DNA does not replicate, the chromosomes condense
again.
6. Metaphase 2. The chromosomes line up on the
equators of the new cells.
7. Anaphase 2. The chromatids separate, one going to
each pole.
8. Telophase 2. The chromosomes gather into nuclei and
the cells divide again.
The whole process of meiosis gives rise to four haploid
gametes, each with a different assortment of chromosomes
and, due to recombination, a different combination of genes
ch02.indd 71
from the chromosomes of the parent. When combined with

a haploid gamete from the other parent they will form part of
a unique individual, who inherits 50% of their genes from
each parent.
DNA REPLICATION
During interphase, which precedes both mitosis and meiosis,
the entire DNA of each cell is copied. This process is a
semi-conservative one in which a new complementary
strand is formed against each of the two parent strands.
The replicated strands will contain one parent strand and
one daughter strand, so only half of the replicated DNA is
new at each cycle of DNA synthesis.
In order to be copied, the double-stranded DNA is
unwound by the action of a number of enzymes, including
helicases, which separate the strands at a position called the
origin of replication. In eukaryotes there are many of these
replication sites. In order to prevent the strand reassembling
during replication, single-stranded binding proteins bind to
each of the separate strands. Replication can proceed in
both directions, producing two replication forks that travel
away from the origin as DNA is unwound and replicated
(Fig. 2.41) . An enzyme called RNA primase, which, with
other proteins, forms the primosome, makes a short strand
of complementary RNA that acts as a primer for the manu
facture of DNA.
The enzyme responsible for the manufacture of new
strands is DNA polymerase III and, like RNA polymerase, it
reads the parent strand only in the 3 to 5 direction, so new
nucleotides are only added to the 3 end of the new strand.
This means that only one of the parent strands, the 3-5
strand, called the leading strand, can be made in one con
tinuous stretch, starting at the 5 end. The other strand, the
lagging strand, is made by the synthesis of multiple short
stretches of about 100200 base pairs long, called Okazaki
fragments. The enzyme DNA polymerase I then replaces the
RNA primer with DNA, and the fragments are then joined
together by DNA ligase at the points where they meet.
DNA checking and repair

DNA polymerase III also controls the accuracy of replication
by checking that the base pair on the original strand and the
new strand are complementary. If they are not, then the
incorrect nucleotide is excised and replaced. This leads to a
2/26/2009 6:48:48 PM

Clinical box 2.14 AZT prevents replication of the
HIV virus because HIV does not
proofread its DNA
The human immunodeficiency virus (HIV) is an RNA virus that
uses the enzyme reverse transcriptase, packaged in the viral
particles, to reverse copy its RNA into DNA, which is then
inserted into the host genome. Reverse transcriptase lacks the
proofreading functions of DNA polymerase, a property which is
exploited in the use of the drug AZT (azidothymidine). AZT is a
nucleoside and is metabolised to an analogue of thymidine
triphosphate, azido thymidine triphosphate. The latter is
incorporated into the elongating DNA chain by reverse
transcriptase and is not corrected. Because the azido group
cannot form a phosphodiester bond with the next nucleotide,
this prevents further elongation of the chain, blocking viral
replication.
very low error rate in replication (Clinical box 2.14). It is esti

mated that the error rate before this proofreading is about
1 in 10000, but with the replacement of wrong nucleotides
this falls to 1 in 1081012. However, this still means that many
genes will contain errors that could be deleterious to the
daughter cell.
Damage to DNA
The DNA of a cell not only contains copying errors but is
continuously being subjected to damage by high-energy
radiation, mutagenic chemicals (particularly reactive oxygen
intermediates) and spontaneous chemical reactions. These
different processes have been estimated to produce up to
60000 modifications per day. Many of these modifications
may produce no discernible effect, possibly because they
occur in non-essential DNA or do not change the activity of
the gene product. However, some may cause the death of
the individual cell. In multicellular organisms with their many
other cells, this can be relatively unimportant. However, more
damagingly, mutations in gametes can lead to damage in
future offspring. This occurs particularly in ova because the
ova are all formed in the early embryo and can therefore
accumulate damage throughout their life from before birth
until the menopause. Mutations in somatic cells can lead to
cancers, with the uncontrolled proliferation of cells.
DNA repair
Two major mechanisms repair DNA continually. They have
been studied extensively in bacteria, but similar mechanisms
are thought to exist in eukaryotes:
Small lesions are repaired by DNA glycosylase
enzymes that check the DNA for mispaired bases,
chemically modified bases, or strands with excess
bases. These are then excised and replaced.
Larger lesions are repaired by removing the section of
DNA containing the damaged, using DNA helicase, and
then replacing it with a new section made by DNA
polymerase, with the new fragment being attached by
DNA ligase.

CELLULAR AGEING AND CELL DEATH

At the same time that new cells are being created, old cells
die. Under normal conditions the adult organism remains a
constant size because cell birth and cell death are balanced.
ch02.indd 72
Clinical box 2.15 Xeroderma pigmentosum

The rare genetic disorder, xeroderma pigmentosum (XP) is
characterised by sensitivity to sunlight and the occurrence of
skin cancers. UV light tends to form pyrimidine dimers that
cannot pair properly with purines during DNA replication,
resulting in basepair substitutions that may result in skin
cancer. Enzymes such as the helicases, endo- and exonucleases, polymerases and ligases normally correct these
pyrimidine dimers. A mutation in any of the genes for these
repair enzymes can produce XP. In the germ line these
mutations are transmitted from one generation to the next. The
deficient repair also leads to persistence of mutations in the
skin cells, which are not inherited but which result in skin cell
tumour development.
Hayflick limit
When cells are removed from the body and placed in tissue
culture, they will grow and divide until they cover the surface
of a dish, forming a monolayer. They then cease dividing and
become quiescent. If the cells are removed from the dish and
re-plated at a lower density, they will start dividing again until
they form a new monolayer. However, normal cells do not
continue in this way indefinitely. Depending on their age
when they were removed from the body, cells continue to
divide about 50 times. This is called the Hayflick limit. Cells
derived from an adult have already divided a number of
times, so they will not divide as many times as those removed
from an embryo. For this reason, cell cultures are usually
prepared from either fetal or young material. Once they reach
their limit, the cells then become senescent and die.
Telomeres
A key element in determining when a cell dies depends on
both the existence of regions of DNA sequences called telomeres and the activity of the enzyme telomerase, which
consists of a combination of protein and RNA. Found at the
end of each chromosome, telomeres are regions of singlestranded DNA composed of repeated nucleotide sequences.
In human telomeres the sequence TTAGGG can be repeated
up to 100 times. The telomeres prevent enzymes that nor
mally join broken ends of DNA from linking the chromosomes
together and allow the chromosome to be copied all the way
to the end. Without the telomeres, the lagging strand would
not be copied to the end, because DNA polymerase cannot
copy the strand underneath the last RNA primer.
Somatic cells do not usually have telomerase activity, so
at each mitotic cell division the telomeres are shortened due
to the failure of DNA polymerase to copy the chromosomal
ends. When the telomeres become too short the chromo
somes tend to fuse together and the cell dies. In contrast, in
germ cells, and in some stem cells, the enzyme telomerase
extends the 3 end of DNA, by a form of reverse transcription:
the RNA template of the enzyme is copied into DNA which
is added to the end of the chromosome, thus maintaining its
length.
The action of telomerase in maintaining cell viability is
shown by cancer cells which are often immortal; that is they
continue to divide beyond the Hayflick limit, seemingly indefi
nitely. These cells have regained active telomerase activity,
maintaining the telomere length. Interestingly, in individuals
with premature ageing diseases, the telomeres are often
found to be unusually short.
2/26/2009 6:48:48 PM
Mechanisms of cell death
Normal cell
Cells can die in two ways:

Necrosis
Apoptosis.
These two mechanisms occur under different conditions

and have very different consequences.
Oxygen toxicity
While oxygen is essential for life on earth, because of
its reactivity, it can also damage cells. Oxygen can react
with a wide range of metabolites via reactions with superoxide anions. First, molecular oxygen is reduced to the super
oxide anion:
O 2 Oi 2 This is a type of free radical (denoted by the dot on the
right) as it has an unpaired electron in its outer shell, making
it highly reactive. Oxygen radicals can react with almost any
biological molecule. They react with lipids to form peroxides,
a process called peroxidation. Most damagingly they react
with DNA, deleterious which can produce mutations that may
be. They can also damage proteins and carbohydrates and
they are particularly important in the development of the
arterial disease, atherosclerosis. This is due to the fact that
oxidised LDLs cannot be taken up in the liver, but are scav
enged by macrophages, which go on to form the foam cells
that are central to the formation of atherosclerotic plaques
(see Ch. 11).
Occasionally, free radical generation can be beneficial
when macrophages use the production of oxygen free radi
cals to kill invading bacteria. NADPH (nicotinamide adenos
ine dinucleotide phosphate (reduced form)) is used to convert
oxygen to superoxide anions. These are then converted to
hydrogen peroxide (H2O2) by the enzyme superoxide dis
mutase and also, with the addition of chloride, to hypochlo
rous acid (HOCl). These compounds are released by the
macrophage in order to degrade bacterial cells. The con
sumption of oxygen required to produce these molecules
results in a respiratory burst that is associated with phago
cytosis (see Ch. 6).
Antioxidants
Damage caused by free radicals can be prevented by the
action of a number of protective compounds, called antioxidants. Many of these compounds, such as vitamin E, form
stable radicals which do not react. An important antioxidant
is glutathione, which is especially important in red blood
cells. Because they carry large amounts of oxygen, red cells
are particularly prone to damage. This is shown by the
appearance of methaemoglobin, an inactive oxidised form of
haemoglobin, which cannot transport oxygen. Glutathione
maintains the proteins and enzymes in a reduced state, par
ticularly the sulphydryl groups, and is also important in
reconverting peroxylipids and proteins, using the enzyme
glutathione reductase. A cofactor for this enzyme is sele
nium, which is required in trace amounts in the diet. Other
antioxidants include vitamin C and -carotene, present in
fruit and vegetables (see Ch. 16).
ch02.indd 73
Apoptotic
bodies
Fig. 2.42 Apoptotic cell death.

Necrosis
When cells are damaged by an acute injury, such as when
they are starved of oxygen by blockage of a blood vessel, as
in a stroke, or poisoned by toxic substances, they die by
necrosis, or accidental cell death. The cells swell and burst,
releasing their cytoplasmic and nuclear contents into the
extracellular space, triggering an inflammatory response.
While this response is a necessary defence mechanism its
effects can also be life-threatening (see Ch. 6).
Apoptosis
Apoptosis, also called programmed cell death (Fig. 2.42), is
the type of cell death that occurs during the normal turnover
of cells in the adult and also under a number of other circum
stances in apparently healthy cells:

Elimination of excess neurons during the development

of the nervous system
During development of organs, such as loss of webbing
between the fingers in the human embryo
Death after a fixed time, such as in the epithelial lining
of the gastrointestinal tract, or in the skin keratinocytes
Shedding of the endometrium during the menstrual
cycle
Death of lymphocytes during their maturation in the
thymus in order to prevent autoimmune reactions (see
Ch. 6).
Apoptosis also occurs when cells are damaged by chronic

injury through viruses, toxins and genetic mutations, but
unlike necrotic cell death, this does not produce a potentially
harmful inflammation.
Cells undergoing apoptosis first lose contact with their
neighbours and shrink. The nuclear chromatin condenses
against the nuclear membrane. The cell then breaks up into
several membrane-bound fragments called apoptotic bodies,
2/26/2009 6:48:48 PM

which are phagocytosed by macrophages or shed from epi
thelial surfaces. Importantly, the cell contents are not released
and there is no inflammatory response.
Triggers for apoptosis

Two main interrelated pathways trigger apoptosis in cells:
the Fas pathway: this involves the expression of cell
surface receptors, called death receptors, which bind
a ligand called Fas. This may be a paracrine or an
autocrine signal. This activates an enzyme, caspase-8,
which in turn activates other caspases (a family of
intracellular proteases) that induce the breakdown of
chromatin.
the Bax pathway: this also involves caspase-8, which
activates the channel protein, Bax. Bax is inserted into
the mitochondrial membrane, allowing cytochrome c to
be released. ATP synthesis in the mitochondria is thus
prevented and cytochrome c also activates other
protease enzymes, which then break down the
intracellular structures.

KEY METABOLIC PATHWAYS

Chemical reactions in the body consist of catabolic reactions, which break down large precursor molecules and
release energy, and anabolic reactions, which use energy in
order to build up large molecules. ATP acts as a form of
energy storage, to be consumed to fuel exergonic or energyrequiring reactions. The metabolic pathways are a highly
integrated network of chemical reactions occurring within
cells. They rely on the energy supplied by the breakdown of
large molecules that may be stored in the body, mainly as
fats or carbohydrates, but which ultimately are derived from
ingested nutrients.
As well as ATP, which transfers phosphate groups from

one molecule to another, a number of other coenzymes
are involved in metabolism and also transfer groups between
molecules. Some of these are given in Table 2.24.
In summary:
1. Carbohydrates and fats are broken down and converted
into glucose and fatty acids, respectively.
2. These are used as substrates in the production of ATP
through a series of pathways that converge on a
common intermediate, acetyl coenzyme A (acetyl CoA)
(Fig. 2.43).
3. Acetyl CoA is further metabolised by a common
pathway called the tricarboxylic acid cycle (TCA), also
known as the citric acid cycle, or Krebs cycle, after
Sir Hans Krebs who first described it in 1937. Under
conditions of decreased carbohydrate and fat intake,
proteins can be broken down into amino acids, and
these can also be metabolised by the same reactions.
4. The reduced coenzymes produced by the TCA cycle are
then used in the conversion of ADP to ATP in a series
of reactions called the electron transport chain. These
chain reactions are all driven by the final step, where
oxygen is required to oxidise cytochrome a.
5. The electron transport chain produces a gradient of
protons across the mitochondrial inner membrane. This
proton gradient is finally dissipated in the conversion of
ADP to ATP by the enzyme ATP synthase in a process
called oxidative phosphorylation.
Glucose
Glycolysis
Amino
acids
Fatty acids
-oxidation
Pyruvate
Information box 2.20 Cancer cells do not

undergo apoptosis
Amino
acids
Acetyl CoA
The protein p53 is expressed by cells in response to

damage to DNA. If the damage is slight, p53 induces the
cells to delay entry to the S phase of the cell cycle, until
the DNA has been repaired. However, if the damage is too
great, then p53 can trigger apoptosis.
A link to cancer is the observation that cancer cells do
not respond to the normal signals that induce apoptosis,
and it has been shown that about 50% of all human
cancers have non-functional p53 genes, hence another
name for p53, the tumour-suppressor gene. p53 also
controls the expression of the Fas and Bax genes. The lack
of p53 expression in cancer cells that have been exposed
to ionising radiation during radiotherapy results in the
survival of these cells, which can then continue to
proliferate.
TCA
cycle
CO2
NADH
FADH2
Electron transport chain
and oxidative phosphorylation
O2
ADP
ATP
H2O
Fig. 2.43 Metabolic pathways to produce acetyl CoA.
Table 2.24 Some coenzymes involved in metabolism
ch02.indd 74
Coenzyme
Group transferred
Oxidised
Reduced
Nicotinamide adenine dinucleotide (NAD)
2H
NAD+
NADH + H+
Nicotinamide adenine dinucleotide phosphate (NADP)
2H
NADP+
NADPH + H+
Flavin adenine dinucleotide (FAD)
2H
FAD
FADH + H+
Coenzyme A (CoA)
Acyl groups (CH3(CH2)nCO), for example, the acetyl

group (CH3CO, where n=0)
CoA
Acetyl CoA
2/26/2009 6:48:49 PM
Key metabolic pathways 75

Glucose
Hexokinase
ATP
ADP
Glucose-6-phosphate (
ATP
glycogen)
glucose transporters in the cell membrane. Glucokinase

has a lower affinity for glucose than hexokinase and, unlike
hexokinase, is not inhibited by glucose-6-phosphate. This
produces two effects:

Fructose-6-phosphate
ATP
Phosphofructokinase
ADP
ATP
Fructose-1, 6-biphosphate
Dihydroxyacetone phosphate
Glyceraldehyde-3-phosphate) (x2)
NAD+
+2NADH
NADH
1, 3-bisphosphoglycerate
ADP
+2ATP
ATP
3-phosphoglycerate
2-phosphoglycerate
Phosphenolpyruvate
ADP
Pyruvate kinase
ATP
In some types of diabetes there are mutations of the

glucokinase gene, which may prevent glucose being trapped
inside the pancreatic -cells. This then prevents glucose-6phosphate levels rising sufficiently inside the cells, and
patients secrete lower amounts of insulin than normal. Red
blood cells lack mitochondria, which contain the enzymes
necessary for further metabolism of pyruvate, so they obtain
all their energy from anaerobic glycolysis.
+2ATP
Pyruvate
Fig. 2.44 Steps in glycolysis.
GLYCOLYSIS
Glycolysis occurs in the cytosol of cells and, via a series of
enzyme-linked reactions, converts the 6-carbon glucose
molecule into two molecules of the 3-carbon pyruvate
(Fig. 2.44). Some of these reactions use ATP (which is
converted to ADP) and others convert ADP to ATP. During
glycolysis there is a net gain of two molecules of ATP for
each molecule of glucose; however, two molecules of NAD
are also reduced, producing two molecules of NADH. None
of these reactions require the presence of oxygen and
therefore they can operate under anaerobic conditions.
However, unless the NADH can be regenerated, the reactions
will stop. NADH regeneration normally occurs by the transfer
of the NADH to the mitochondria where it can be
reoxidised.
In strenuous exercise, the rate of this reoxidation is insuf
ficient and, in order to continue the breakdown of glucose,
NADH is oxidised by the reduction of pyruvate to lactate. This
is exported to the liver, where it can be used to resynthesise
glucose (see Ch. 3). Many of the intermediate molecules
of glycolysis can also be used in the synthesis of other
biomolecules.
Hexokinase and glucokinase

The first step in glycolysis is the phosphorylation of glucose
to glucose-6-phosphate, either by the enzyme hexokinase,
present in all tissues, or glucokinase, which only occurs
in the liver or in the insulin-secreting pancreatic -cells.
This important reaction traps the glucose within the cell as
glucose-6-phosphate, which is not a substrate for the
ch02.indd 75
The liver can take up large amounts of glucose from

the glucose-rich blood arriving via the hepatic portal
vein from the intestine after a carbohydrate-rich
meal.
In pancreatic -cells, because the enzyme does not
saturate at normal levels of blood glucose, the
concentration of glucose inside the cells continues to
rise even after the ingestion of large amounts of
glucose. As the secretion of insulin is linked to the
amount of glucose-6-phophate inside the cells, this
ensures that, after a large meal, sufficient insulin is
released.
Control of glycolysis
The rate of glycolysis in red blood cells is regulated by con
trolling the activity of three enzymes in the glycolytic pathway.
These are:

Hexokinase
Phosphofructokinase (PFK-1), which converts
fructose-6-phosphate to fructose-1,6-bisphosphate
Pyruvate kinase, which catalyses the last step in
glycolysis, which produces pyruvate.

All these three enzymes catalyse irreversible reactions

(although some reactions can be reversed by other enzymes)
which, because of their low activity levels (low Vmax) relative
to other glycolytic enzymes, are rate limiting steps.
Hexokinase
Hexokinase is inhibited by its product, glucose-6-phosphate,
and has the lowest activity of all the glycolytic enzymes.
PFK-1
The dominant role in the control of glycolysis is the allosteric
regulation of PFK-1. Binding of ATP to the enzyme lowers
its affinity for the substrate (i.e. it increases Km). This enzyme
is also stimulated by ADP and AMP, so its overall activity
depends on the ratio between ATP and (ADP + AMP)
concentrations.
Pyruvate kinase
The third enzyme, pyruvate kinase, is activated by the product
of the PFK-1 reaction, fructose-1,6-bisphosphate. This type
of regulation, known as feed-forward activation, ensures
that intermediates of glycolysis do not accumulate. Glycoly
sis is inhibited by other metabolites such as pyruvate and
citrate (which is an intermediate in the TCA cycle; see
below).
2/26/2009 6:48:49 PM

Control mechanisms in other cells
All tissues except the brain and red blood cells take up free
fatty acids from the blood and use them in the production
of acetyl CoA (see Ch. 3). Fats are energy-rich foods as
they produce more ATP per mole when oxidised than
glucose, owing to the large number of H atoms in the fat
molecules.
Link reaction
The carnitine shuttle
Pyruvate produced by glycolysis is transported into the mito

chondria where the rest of energy metabolism takes place.
The oxidation of pyruvate to acetyl CoA occurs via a link
reaction catalysed by the enzyme complex pyruvate dehydrogenase, which requires the coenzyme thiamine pyro
phosphate (derived from vitamin B1) as well as other
coenzymes. During this reaction, for each molecule of pyru
vate a single carbon is lost as CO2 and a molecule of NADH
produced (Fig. 2.45). As well as being converted to acetyl
CoA and lactate, pyruvate can also be converted to alanine,
which can be used in protein synthesis, or to oxaloacetate,
to be used by the liver to synthesise glucose.
Link reaction
Pyruvate
CoA + NAD2
Pyruvate dehydrogenase
NADH + CO2
Acetyl CoA
Fate of pyruvate
Pyruvate
CH3CO COO
(Acetyl)
Transferred
to CoA
Released
as CO2
Fig. 2.45 Link reaction between glycolysis and the

TCA cycle.
Clinical box 2.16 Vitamin B1 deficiency impairs

glucose metabolism
Vitamin B1 is the source of an essential coenzyme involved in
carbohydrate metabolism. The body stores very little vitamin B1
(thiamine) and so it has to be ingested at regular intervals.
Thiamine is present in the husks of rice, in wheat germ and in
seeds. Thiamine is phosphorylated in the tissues to produce
both the coenzyme, thiamine pyrophosphate (also known as
thiamine diphosphate), and thiamine triphosphate, which has a
role in nerve conduction. In thiamine deficiency there are raised
levels of pyruvate, which will inhibit glycolysis. This can lead to
the accumulation of glucose in the blood. The accumulation of
both pyruvic acid and lactic acid causes acidosis which, in
severe cases, can lead to coma and death.
The commonest form of thiamine deficiency, beriberi, occurs
in populations dependent on rice diets where, if the rice is
polished, thiamine is removed. Alcohol inhibits the absorption of
thiamine in the intestines, so deficiency conditions are relatively
common in alcoholics. WernickeKorsakoffs syndrome is the
result of thiamine deficiency associated with the combination of
chronic misuse of alcohol and malnutrition.
ch02.indd 76
THE METABOLISM OF FATTY ACIDS
The control mechanisms described above occur in red blood

cells, which seem to have relatively simple requirements, and
their metabolism remains fairly constant. In other tissues
such muscle and liver, however, where energy requirements
fluctuate more, the control of these key enzymes is more
complicated, involving other allosteric modulators and cova
lent modification, such as phosphorylation.
The first step involves the conversion of fatty acids to a CoA

derivative (acyl CoA) as they enter the cell, which prevents
the fatty acids dissolving cell membranes. While small acyl
CoA molecules derived from fatty acids with fewer than
about 12 carbons can passively diffuse into mitochondria,
larger acyl CoA molecules are transported across the mito
chondrial membranes by a shuttle mechanism. This involves
their conversion to acylcarnitine, with the replacement of the
CoA by carnitine, leaving CoA in the cytoplasm. The resulting
acylcarnitine is transported across both the outer and inner
membranes, before being converted back to acyl CoA and
carnitine in the matrix. The carnitine is then returned to the
cytosol where it can pick up another acyl CoA. As the trans
porters on both membranes use antiport mechanisms, the
ability to transport fatty acids into the mitochondrial matrix
is regulated by the level of free CoA in the matrix. If fatty acid
levels are high then most of the CoA will be acylated and no
further transport can occur.
-Oxidation
The catabolism of acyl CoA molecules is carried out in
the mitochondrial matrix by a process called b-oxidation
(Fig. 2.46). This occurs via the stepwise removal of two
carbon units to produce acetyl CoA. This process not only
produces large amounts of acetyl CoA, but each step
produces one NADH and one FADH2. For each fatty
acid of n carbon atoms, there are n/2 molecules of
acetyl CoA produced and (n/2 1) of each of NADH and
FADH2.
Each cycle is a series of four steps:
1. An oxidation reaction of the -carbon (hence the name
-oxidation) to form a double bond (Fig. 2.46 step 1),
yielding one FADH2.
2. This double bond is then hydrated to produce a
hydroxyl group (Step 2).
3. The hydroxyl group is then oxidised (Step 3), yielding
one NADH.
4. This is then cleaved by a thiolase enzyme to give a
molecule of acyl CoA that is shorter by two carbon
atoms and a molecule of acetyl CoA (Step 4).
The enzymes which carry out these four steps are thought
to be closely associated in the membrane so that the product
of each reaction is passed directly to the next enzyme. As a
consequence none of the intermediates can be detected in
the mitochondrial matrix.
There are three different acyl CoA dehydrogenase
enzymes which catalyse step 1, each with specificity for
either short, medium or long chain acyl CoA. Very-long-chain
fatty acids (>20 carbons) are shortened in peroxisomes to
enable their uptake by mitochondria. Peroxisomes can also
carry out -oxidation of fatty acids with the production of
hydrogen peroxide (H2O2), instead of FADH2.
2/26/2009 6:48:50 PM

O
Fatty acyl CoA

R
CH2
CH2
Acetyl CoA
S
FAD
FADH2
Step 1
Oxaloacetate
ADH
O
R
CH
CH
CoA
Citrate synthase
CoA
Citrate
NADH
S
CoA
Malate
NAD+
Isocitrate
NAD+
H2O
Step 2
NADH
+CO2
H2O
OH
R
CH
O
CH2
Step 3
O
R
CH2
Fumarate
S
CoA
Succinate
Fig. 2.47 The TCA cycle.
Succinyl CoA
CoA
TCA CYCLE
O
C
CoASH (coenzyme A)
S
CoA
Thiolase
Fatty acyl CoA
O
CH3
CoA
Acetyl CoA
Fig. 2.46
FAD
NAD+ -Ketoglutarate
+CO2
CoA
GTP NADH
Dehydrogenase
CoA
GDP
NAD+
NADH
Step 4
FADH2
Isocitrate
dehydrogenase
-Oxidation of fatty acids. R = CH3-(CH2)n
The TCA cycle starts with the formation of citrate from a

condensation reaction between acetyl CoA and the fourcarbon oxaloacetate. This releases the CoA, which can then
react with a further molecule of pyruvate. This series of reac
tions is called a cycle rather than a pathway because it
eventually produces oxaloacetate which can react with
another molecule of acetyl CoA. However, during these reac
tions, for each molecule of acetyl CoA, three molecules of
NADH, one of FADH2 and one molecule of GTP are produced
(Fig. 2.47).
Control of the TCA cycle
In the case of unsaturated fatty acids, enzymes alter the

position and shape of the double bonds.
With odd numbers of carbons the last reaction
produces propionyl CoA (which contains an extra CH2
group), which is converted to the TCA cycle
intermediate, succinyl CoA.
The TCA cycle is regulated by three enzymes. Citrate synthase, which catalyses the condensation reaction between
acetyl CoA and oxaloacetate, is sensitive to the availability
of its substrates, particularly oxaloacetate. This enzyme is
also inhibited by ATP and allosterically activated by ADP. In
this way it is sensitive to the energy state of the cell. The
other key regulatory enzymes, isocitrate dehydrogenase
and -ketoglutarate dehydrogenase, are both regulated
by the levels of NAD+ and NADH. In this way the cycle can
respond to the cells need for energy.
Ketones
Uses for TCA cycle intermediates
Fatty acids that are unsaturated or have uneven numbers

of carbons can also be converted to acetyl CoA, but there
are additional steps involved.

In the liver any surplus acetyl CoA produced from fatty acid
breakdown can be converted to ketones bodies or ketones,
which can be exported to other organs. Ketone bodies
consist of acetoacetate, -hydroxy butyrate and acetone,
which are water soluble. This type of metabolism is important
for many tissues (except the liver) in fasting, starvation and
in diabetes mellitus. However, high levels of acetoacetate
and -hydroxy butyrate can cause an increase in the acidity
of the blood, a type of metabolic acidosis (see Ch. 1).
During starvation, brain tissue may acquire 50% of its energy
from ketone bodies. This reduces the demand for glucose
and also thereby reduces the need for glucose production
from other sources (see Ch. 3), particularly from the break
down of body protein a potentially survival-enhancing
metabolic switch.
ch02.indd 77
As well as providing reduced coenzymes for the electron

transport chain, many of the intermediates of the TCA cycle
can be used as substrates for other anabolic pathways
(Fig. 2.48).
Examples are:

Citrate can be used as a substrate for fatty acid

synthesis
Oxaloacetate can be converted to aspartate from
which it can be used in many ways, including the
manufacture of amino acids, nucleic acids and in the
urea cycle, or as the starting point for the synthesis of
glucose (gluconeogenesis)
succinyl CoA can be used as the starting point for the
synthesis of haem.
2/26/2009 6:48:50 PM

Alanine
Glucose
Pyruvate
Aspartate
Urea cycle
Fatty acids
Acetyl CoA
Table 2.25 Glucogenic and ketogenic amino acids
Ketone bodies
Type
Amino acids
Glucogenic only
Alanine, arginine, asparagine,

aspartate, cysteine, glutamate,
glutamine, glycine, histidine,
methionine, proline, serine, valine
Ketogenic only
Leucine, lysine
Both glucogenic and

ketogenic
Isoleucine, phenylalanine,
threonine, tryptophan, tyrosine
Fatty
acids
Oxaloacetate
Malate
Citrate
-Ketoglutarate
Fumarate
genic amino acids give rise to pyruvate or a number of TCA

cycle intermediates.
Glutamate
Succinate
Essential amino acids
Succinyl CoA
Haem
Other
amino acids
Fig. 2.48 TCA cycle intermediates are substrates for

many other reactions.
While many of the 20 amino acids used to manufacture pro

teins can be synthesised in the body there are a number that
cannot. Adults require adequate dietary amounts of eight
amino acids:

AMINO ACID METABOLISM

Proteins are broken down into amino acids (see also Ch. 3).
Many of these are used directly in the production of other
proteins and a number of important metabolites, including
the purines and pyrimidines used in nucleic acid synthesis,
some neurotransmitters, and the haem portion of haemo
globin. Amino acids can also be used to provide energy,
particularly during fasting.
Deamination and transamination

Before amino acids can be used as fuel their amino groups
must be removed, leaving the corresponding -keto acid.
This occurs either by deamination, which directly oxidises
the amino acid to its keto acid with the removal of ammonia,
or transamination, in which the amino group is transferred
to an acceptor, another keto acid, which is thus converted
to an amino acid. For example, the removal of the amino
group from alanine (CH3CH(NH2)COOH) produces pyruvate
(CH3COCOOH). Similarly, the removal of the amino group
from glutamate converts it to -ketoglutarate, one of the TCA
cycle intermediates. In this way amino acid derivatives can
be integrated directly into energy metabolism.
Urea cycle
Ammonia is highly toxic, so it is immediately converted to
glutamine or alanine. These are eventually removed, mainly
by conversion into urea, carried out by the urea cycle in the
liver. The urea is subsequently excreted in the urine.
Glucogenic and ketogenic

amino acids
When the carbon skeletons derived from amino acids are
catabolised, depending on metabolic requirements, they can
be converted into glucose or ketone bodies. Different amino
acids yield different products, so they are called glucogenic,
or ketogenic, or both (Table 2.25). Ketogenic amino acids
can yield either acetyl CoA or acetoacetyl CoA, while gluco
ch02.indd 78
Isoleucine
Leucine
Lysine
Methionine
Phenylalanine
Threonine
Tryptophan
Valine.
Infants also require histidine and arginine as well because

the amounts they can synthesise may not be enough to
support their growth rate (see Ch. 16). Cysteine and tyrosine
can only be made from the essential amino acids methionine
and phenylalanine, respectively, so if they are not present
in the diet, more methionine and phenylalanine must be
ingested.
ELECTRON TRANSPORT CHAIN AND

OXIDATIVE PHOSPHORYLATION
The final part of the transformation of glucose and other
energy supplies into a source of energy usable by metabolic
reactions occurs on the inner membrane of the mitochon
drion. The enzymes responsible are mostly integral to the
inner membrane and are arranged as a sequence of protein
complexes called the electron transport chain.
Electron transport chain

Starting with the reduced coenzymes, electrons are passed
from complex to complex, with the complexes acting as
proton pumps (Fig. 2.49). The energy required to pump
protons, against their concentration gradient, into the space
between the inner and outer mitochondrial membranes is
obtained from the free energy released as the electrons move
from complex to complex, down a gradient of redox potentials. The final electron acceptor in the chain is oxygen,
which combines with hydrogen to give water.
The overall reaction is therefore: glucose plus oxygen
gives carbon dioxide and water.
C6H12 O6 + 6O2 6CO2 + 6H2 O plus 38 ADP molecules
converted to ATP
2/26/2009 6:48:51 PM

ATP synthase
ATPsynthase
complex
complex
Outer membrane
H+
H+
Inter-membrane
space
Inner membrane
ATP: ADP
translocase
H+
Cristae
CO2
Pi + ADP
H+
ATP
Reduced coenzymes
FADH2
Matrix
H+
H2O
H+
H+
H+
1
2
O2
e
H+
IV
III
H+
H+
e
e
NADH
H+
ADP
Fuel
oxidation
e
CO2
TCA
cycle
ATP
OH
I
H+
Pi
Proton pumps for

electron transport system
(complexes I, III and IV)
Fig. 2.49 Energy transfer in the mitochondrion. Adapted with permission from Baynes J, Dominiczak M 2005 Medical
biochemistry, 2nd edn. Elsevier Mosby, Edinburgh.
Electrons from NADH can combine with complex I, while

those from FADH2 are transferred via complex II. These
pathways converge at ubiquinone (coenzyme Q10). The elec
trons are subsequently passed to complex III and then via
cytochrome c to complex IV, from where the electrons are
finally accepted by oxygen. Each of complexes I, III and IV
acts as a proton pump, increasing the concentration of H+
ions in the intermembrane space.
Redox potentials drive metabolic reactions

The redox potential is the force that transfers electrons from
one compound to another in oxidationreduction (or redox)
reactions. The gain of electrons is a reduction, and the loss
an oxidation. When a reducing agent loses electrons, energy
is transferred to the reduced product. Each molecule in the
electron transport chain has a lower potential energy (a lower
redox potential) than its predecessor, resulting in progressive
release of energy. This energy is transferred to the gradient
of hydrogen ions across the inner mitochondrial membrane.
Because hydrogen is concentrated in the intermembrane
space and not at equilibrium, the energy released by the
redox reactions is stored as the potential energy of the pH
gradient.
Chemiosmotic theory of
oxidative phosphorylation
The chemiosmotic theory was devised to explain how the
proton gradient is used to produce ATP. Inserted into the
inner mitochondrial membrane, the multimeric protein ATP
synthase, also known as complex V, consists of two major
components, known as F0 and F1. Hydrogen ions flow down
their electrochemical gradient through F0,, which is a trans
membrane protein with a central channel.
Attached to F0 via a stalk region is the F1 protein which
converts ADP to ATP in a process called oxidative phosphorylation. Each pair of electrons passing along the elec
tron transport chain reduces one atom of oxygen (O2), and
each of the three proton pumps moves enough H+ to power
the production of 1 molecule of ATP.
ch02.indd 79
Table 2.26 Summary of ATP generated during the

complete oxidation of glucose
Stage
Net change in coenzymes

(per mole glucose
entering glycolysis)
Glycolysis
2 ATP
2 NADH
2
6
Link reaction
2 NADH
TCA cycle
2 GTP
6 NADH
2 FADH
2
18
4
ATP equivalents*
Total 38
*During aerobic metabolism 1 NAD = 3 ATP and 1 FAD = 2 ATP.
Thus, when it is fully oxidised, each molecule of NADH

will produce three molecules of ATP, while FADH2 (which
enters via complex II, bypassing the proton pump of complex
I) only produces two. It is therefore possible to calculate
the number of moles of ATP that can be produced by the
metabolism of a mole of glucose (Table 2.26). The exact ratio
between proton pumping and ATP synthesis varies, however,
so 38 moles of ATP per mole glucose is an approximation.
A comparative calculation for the 16-carbon fatty acid,
palmitic acid, gives a total of 129moles of ATP per mole.
This increases by 17moles of ATP for each subsequent
2-carbon increase in fatty acid length. So stearic acid
(C18) metabolism yields 146moles of ATP per mole, which
shows why fats provide so much more energy than
carbohydrates.
Coupling of the electron transport

chain and oxidative phosphorylation
The electron transport chain and oxidative phosphorylation
are tightly coupled. This can be shown in vitro by measuring
oxygen utilisation (a measure of the end point of the electron
transport chain) while controlling ATP synthesis. If ADP is not
added to the reaction then oxygen utilisation stops. When
ADP is included, oxygen use increases until it is all converted
to ATP. Blocking the proton channel of F0 with oligomycin in
2/26/2009 6:48:51 PM
Information box 2.21 Cyanide and other poisons
Studies of the effect of a number of compounds, including

cyanide, on in vitro preparations of mitochondria (usually
from rat liver) have helped to elucidate the sequence of
steps involved in the electron transport chain and oxidative
phosphorylation. At the same time, because of their mode
of action, they can also act as potent poisons in vivo.
In the presence of inhibitors of complex I, such as the
insecticide rotenone, NADH cannot be oxidised but FADH
can. The addition of succinate, which is metabolised in the
TCA cycle to yield FADH, can stimulate the production of
ATP while malate, whose metabolism yields NADH, does
not.
A compound called antimycin A inhibits complex III.
However, while the oxidation of both NADH and FADH are
blocked, the addition of ascorbate (which can directly
reduce cytochrome c, and which normally passes electrons
from complex III to IV) restores ATP production.
Cyanide and carbon monoxide both inhibit complex IV,
as well as binding to haemoglobin. As electrons can no
longer be passed from complex IV to oxygen, the
movement of all electrons stops, and hence both proton
pumping and ATP production cease. In the absence of
aerobic respiration, anaerobic lactate metabolism is
switched on, resulting in a lactic acidosis, which can lead
to death. One antidote is methylene blue, which removes
the inhibition by accepting electrons from complex III;
another is thiosulphate, which converts cyanide to
thiocyanate.
the presence of ADP stops oxygen use, showing that it is the

dissipation of the proton gradient that determines the rate
of the electron transport chain. This is because the electron
transport chain can only generate a gradient equivalent to
two pH units. If the gradient exceeds this, then the proton
pumps cannot generate sufficient energy to continue.
cells that are adapted to perform a specific function. Organs

are where more than one tissue forms a structural unit with
a particular function.
ORIGINS OF CELLS
During the very early development of the embryo, by 16 days,
the embryo has developed three layers of cells, called (from
the surface inwards):

Ectoderm
Mesoderm
Endoderm.

It is from these three germ cell layers that all cell types
form. Some cells, such as epithelial cells, form from all three
germ cell layers. However, other cells are derived from single
layers. For example, nervous tissue is derived from ecto
derm, muscle and connective tissue mainly from mesoderm,
and most mucosae from endoderm.
In the adult only some cell types remain highly mitotic,
such as epithelial and blood-forming cells. Others, such as
nervous tissue, rarely divide. Some cells are produced by
division of mature differentiated cells, but many new cells are
derived from relatively undifferentiated stem cells. Stem cells
have now been found in many different tissues. Of great
interest is the possibility of using stem cells from easily
accessible sources, such as the blood, to produce replace
ment cells in organs such as the brain where stem cells are
rarer (see Information box 2.22).
BASIC TISSUE TYPES

There are four primary tissue types in the body. These are:

Epithelial tissues
Connective tissues (including blood)
Muscle tissues
Nervous tissues.

Uncoupling of the electron

transport chain
Compounds called uncouplers, such as 2,4-dinitrophenol
(DNP), do not affect the electron transport chain but act in a
way that allows the proton gradient to dissipate, without the
production of ATP. In the presence of DNP, oxygen utilisation
increases in an attempt to restore the proton gradient.
Uncoupling of the proton gradient allows the energy stored
in the gradient to be dissipated as heat.
This mechanism is very important in newborn babies as
a source of heat. Neonates do not shiver when they get cold,
possibly because their nervous systems are not sufficiently
developed to coordinate the muscular activity required. So,
in specialised areas of adipose tissue called brown fat (so
called because of the high number of mitochondria), an
uncoupling protein called thermogenin allows the electron
transport chain to be used under controlled conditions to
generate heat rather than ATP.
TISSUES AND ORGANS

Although a typical cell can be described and the function
of its organelles explained, almost all the cells in the body
show some specialisation, and most cells are organised into
tissues and organs. Tissues are defined as a collection of
ch02.indd 80
Epithelial tissues
Epithelial tissues, also called epithelium, are sheets of cells
which cover or line body surfaces and form secretory glands.
They are derived from all three of the germ cell layers found
in the early embryo and have a variety of functions, which
include secretion, absorption, protection and transport. They
are characterised by a high cell density (a high cellularity).
Epithelial tissues have a well-defined polarity with the basal
end of the cell, which sits on a basement membrane, and
a luminal end, which normally faces the surface, such as the
intestinal lumen or the bloodstream. They are joined in sheets
by intercellular junctions. The basement membrane provides
mechanical support and attachment for the epithelial cell. In
the kidney, the basal lamina also provides a barrier for the
filtration of blood components during the formation of urine
(see Ch. 14). Many epithelial cells have specialisations of
their apical surfaces, such as microvilli and cilia.
Classification of epithelial cells

Epithelial tissues, which form linings, are usually classified
according to three criteria (Table 2.27 and Fig. 2.50):

How the sheets are made up. Epithelial sheets may be

made up of either a single layer of cells, a simple
2/26/2009 6:48:51 PM
Tissues and organs 81

Table 2.27 Types of lining epithelium
Type
Cell shape
Examples
Function
Simple squamous
Flattened
Pulmonary alveoli; loop of Henle;

parietal layer of Bowmans capsule;
endothelium of blood vessels and
lymphatic vessels; mesothelium of
pleural and peritoneal cavities
Limiting membrane; fluid

transport; gaseous exchange;
lubrication; reducing friction;
lining membrane
Simple cuboidal
Cuboidal
Ducts of many glands; covering of

ovary; kidney tubules
Secretion; absorption; protection
Simple columnar
Columnar
Oviducts; uterus; small bronchi; much

of digestive tract; gall bladder; large
ducts of some glands
Transportation; absorption;
secretion; protection
Pseudostratified columnar
While all cells are in contact

with the basement
membrane they do not all
reach the luminal surface.
Those that do are columnar
Respiratory epithelium; most of

trachea; primary bronchi; epididymis;
vas deferens; large secretory ducts
Secretion; absorption; lubrication;

protection; transportation
Stratified squamous
(non-keratinised)
Flattened (with nuclei)
Mouth; oesophagus; vagina
Protection; secretion
Stratified squamous
(keratinised)
Flattened (without nuclei)
Epidermis of skin
Protection
Stratified cuboidal
Cuboidal
Ducts of sweat glands
Absorption; secretion
Stratified columnar
Columnar
Conjunctiva of eye; some large

secretory ducts; portions of male
urethra
Secretion; absorption; protection
Transitional
Dome-shaped (relaxed)
flattened (distended)
urothelium urinary tract from renal

calyces to urethra
Protection; distension
Simple
Stratified
epithelium, or from several layers, called stratified

epithelium.
The cell shape. The cells may have different shapes:
cuboidal, squamous (flattened), columnar, or transitional
(where the cells vary in shape across the different layers
of the stratified epithelial sheet).
The type of surface specialisation, for example cilia or
keratin.
Glandular epithelial cells

Glandular epithelial cells store and secrete many compounds
such as hormones and enzymes. The most simple type of
glandular epithelium are the unicellular goblet cells, which
secrete mucus into the intestinal lumen. Epithelial cells are
also organised into glands. Glandular epithelia form either
exocrine glands, which are continuous with the body sur
faces and secrete either to the outside of the body or into
luminal spaces via an excretory duct, or endocrine glands,
which are ductless and where the secretions pass directly
into the bloodstream. For example, the sweat glands are
continuous with the skin surfaces. In contrast, endocrine
glands, such as the adrenal gland, surround blood vessels
and secrete hormones into the blood.
Exocrine secretion
Exocrine glands have a secretory portion, where the secre
tory product is released into the lumen of the gland, and an
excretory portion consisting of a duct, by which the secretory
product is transported to the outside. Exocrine glands are
classified as either simple (such as the sweat glands), where
the duct is unbranched, or compound where the secretory
duct is branched and the gland is divided into units called
lobes, which themselves are further subdivided into lobules.
ch02.indd 81
A compound gland, such as the salivary gland, contains

many different types of epithelial cells in different parts of the
gland.
There are three types of secretion:
Merocrine, where the release is by exocytosis
Apocrine, where part of the apical surface is pinched
off with the secretions inside it
Holocrine, where the entire cell disintegrates to release
the stored product.

In the mammary glands, milk proteins are secreted by

merocrine secretion (exocytosis) and milk lipids are secreted
by apocrine secretion. An example of holocrine secretion
occurs in the sebaceous glands where sebum stored in the
cytoplasm is released when the entire cell disintegrates.
Connective and supporting tissues

There are four types of connective and supporting tissues,
all of which are derived from mesenchymal multipotent stem
cells in the mesoderm. Between them they provide the struc
tural, metabolic and defensive support within all tissues and
organs. These are:

Fibrocollagenous tissues
Cartilage, bone and teeth
Adipose tissue
Blood.

ECM
Characteristically, connective tissue contains two compo
nents, the ECM and the cells, which are relatively sparse
when compared to epithelial tissues. The ECM contains
ground substance, made up of three main elements:
2/26/2009 6:48:51 PM
Simple squamous
A
A
Simple cuboidal
B
B
Simple columnar
C
C
Pseudostratified columnar
D
D
Stratified squamous
E
E
Stratified cuboidal
F
F
Transitional
Fig.
ch02.indd 82
Distended
2.50 Epithelial cell types.
Relaxed
2/26/2009 6:48:55 PM

A
Loose or areolar connective tissue

Submucosa of large intestine
Dense connective tissues

Dermis of the skin
D
Reticular tissue
Liver
Tendon
Fig. 2.51 Types of fibrocollagenous tissue.
Structural carbohydrates, the glycosaminoglycans

(GAGs) which form large proteoglycan complexes
Structural proteins such as laminin and fibronectin
Fibres, which are either collagen or elastin; collagen
provides strength while elastin provides elasticity and
support.

Fibrocollagenous tissues
Fibrocollagenous tissues can be divided into three main
types (Fig. 2.51):

Areolar or loose connective tissue, with abundant

ground substance, which gives it a gel-like consistency,
some collagen and elastic fibres, as well as many cell
types.
Dense connective tissue, which has little ground
substance and abundant collagen and a few cells.
These tissues provide mechanical support and tensile
strength. Two types of dense connective tissue are
defined according to the organisation of the collagen
fibres:
Irregular which is found in the dermis of the skin
and the submucosa of the digestive tract
Regular which is found in tendons and ligaments.
This type of tissue is the most dense, with the least
blood supply.
Reticular connective tissue, which only occurs in organs
with a high cellularity, such as the liver, where reticulin
fibres (type III collagen) form a fine network around the
many epithelial cells.
ch02.indd 83
Fibroblasts
A number of different cells are found in fibrocollagenous
connective tissue. Fibroblasts are spindle-shaped cells with
an oval nucleus. They have a well-developed ER and a Golgi
apparatus, which is indicative of their role in the secretion of
ECM components. They produce the mature proteoglycans
of the ECM as well as the precursors of the collagens (tropocollagen) and the elastins (tropoelastin). Fibroblasts are
very active in wound healing, producing new connective
tissue to bridge the wound. Fibroblasts which can contract,
myofibroblasts, are involved in the shrinkage of the scar
tissue.
Other cells in connective tissue

Also present in fibrocollagenous tissue are blood vessels,
small numbers of adipocytes which store fat, stem cells
(mesenchymal cells which can differentiate into a variety of
cell types to replace, repair or grow existing tissues) and a
wide variety of cells of the immune system (see Ch. 6). These
include:

macrophages cells of the immune system which can

engulf (phagocytose) and digest bacteria, cell debris
and other unwanted material.
mast cells cells of the immune system that contain
granules. Some of these contain histamine and other
mediators of inflammation, which are released following
exposure to environmental allergens.
plasma cells these are antibody-secreting mature B
cells of the immune system.
2/26/2009 6:48:58 PM

Cartilage
Cartilage is denser than dense connective tissue, but less
dense than bone. It is solid but flexible and resists compres
sion while allowing diffusion of water through the matrix. It
lacks blood vessels, so all metabolites must be exchanged
by diffusion. The ground substance of cartilage has large
amounts of the GAGs chondroitin sulphate and hyaluronic
acid, bound to a lattice of type II collagen. Growing cartilage
contains chondroblasts, metabolically active cells which
secrete proteins and contain energy reserves in the form of
lipids and glycogen. In adult cartilage the much less active
mature chondrocytes are found in cavities, called lacunae,
in the ECM.
The three types of cartilage reflect the varying amounts
and types of fibres:
Hyaline cartilage contains type II collagen only, forms
most of the embryonic skeleton, and in the adult forms
the sternal part of the ribs and also the cartilage found
in the nose, trachea and larynx. It also covers the ends
of the long bones where it can absorb some of the
compressional stresses.
Elastic cartilage this has more elastic fibres than
hyaline cartilage. It supports the pinna of the external
part of the ear and forms the epiglottis.
Fibrocartilage has less matrix than hyaline cartilage
and also contains type I collagen. It is more
compressible than hyaline cartilage and is found in
areas where there are high pressures, such as the
intervertebral discs and the knee joint.

In post-menopausal women one of the effects of the lack of

the sex steroid hormone, oestrogen, is a reduction in bone
density. This makes the bones fragile and more likely to
fracture. This is due to an increase in the numbers of
osteoclasts leading to an increased bone reabsorption. This
can be reversed in post-menopausal women by hormone
replacement therapy (HRT), plus dietary calcium supplements
and vitamin D if necessary.
Teeth
Teeth are made up of three layers. The external surface, or
crown, of the tooth is covered by enamel, the hardest sub
stance in the body. It is principally made up of calcium
phosphate. The middle layer consists of a mineralised matrix,
called dentine, similar to bone although it does not contain
cells. Inside the dentine is the pulp cavity, which contains
the cells which produce the dentine, the odontoblasts, as
well as the nerves and blood vessels supplying the tooth.
These enter the tooth through the root canal, a narrow
channel at the root of the tooth. The part of the tooth which
is embedded in the jaw bone is covered by a thin layer of a
calcified tissue called cementum, which serves both to
anchor the tooth and protect it.
Adipose tissue
Both hyaline and elastic cartilage are surrounded by a

layer called the perichondrium, which consists of a fibrous
layer that is continuous with the periosteal bone and the
surface of surrounding connective tissue.
While small numbers of adipocytes are found either isolated

or in clumps throughout all loose supporting tissues, in
adipose tissue they form the major cell type. There are two
types of adipose tissue: white and brown. These are both
derived from a common precursor cell, the preadipocyte,
which itself is derived from the mesenchymal stem cell.
Bone
White adipose tissue
Bone is similar in some ways to cartilage, except that the

ECM has more collagen fibres (type I) and the matrix has
become mineralised. This produces a very rigid tissue that
forms the skeleton (see Ch. 9). The ground substance con
tains glycoproteins, which specifically bind calcium, called
osteocalcin. Bone is initially formed by osteoblasts, which
produce the matrix called osteoid; this is followed by the
deposition of calcium phosphate. Crystals of calcium
hydroxyapatite are formed by the addition of hydroxide and
bicarbonate ions to the amorphous calcium phosphate. This
accumulates around the osteoblast, which becomes an
osteocyte.
White adipose tissue contains adipocytes whose entire

cytoplasm is taken up by a single large droplet of fat. The
nucleus is flattened and pushed to one side and the cyto
plasm, which contains mainly mitochondria, exists only as a
ring around the fat droplet. Adipose tissue not only provides
a large energy store, containing about 80% triacylglycerol,
but also acts as a shock absorber and insulator. It is found
throughout the body, particularly under the skin and around
abdominal organs. The amount of adipose tissue varies with
age and gender, typically increasing with age and being
higher in women.
Woven and lamellar bone
Brown adipose tissue is only extensive in newborn mammals

and some hibernating animals. It is highly vascularised and
contains multiple droplets of fat that surround a large central
nucleus. The cells contain large numbers of mitochondria
that metabolise the fat without the production of ATP. This
uncoupling produces heat when the cells are stimulated by
the sympathetic autonomic nervous system. The name
brown fat derives from the colour of the large amount of
cytochromes (electron transport chain molecules) present in
the mitochondria. In adults, brown fat may have a role in
burning off excess fat and preventing obesity.
There are two major types of bone:

1. Woven bone is an immature form, which is relatively
weak and has randomly organised collagen fibres.
Woven bone is produced when osteoid is being formed
rapidly, as in the embryo or during the early repair of a
fracture.
2. Woven bone is then remodelled into lamellar bone,
where the collagen is highly organised into layers, or
lamellae. Lamellar bone has a highly organised
structure, which is highly vascularised.
Mature bone is continually being remodelled due to the
activity of osteoclasts, which break down bone, and oste
oblasts, which replace it.
ch02.indd 84
Clinical box 2.17 Osteoporosis is caused by an

increased reabsorption of bone
Brown adipose tissue
Blood
Although blood does not provide mechanical support or
connections between tissues, it is classified as a connective
2/26/2009 6:48:58 PM

Clinical box 2.18 Understanding the control of
adipose tissue is of interest in
the fight against obesity
People who are above average weight have a higher mortality
than those who weigh less. The most commonly used measure
of obesity is the body mass index (BMI), which is calculated
from the weight (w) in kg and height (h) in m, where BMI = w/h2.
The longest life expectancy is associated with a BMI between
20 and 25, with higher mortality for those underweight (BMI
<20) and overweight (BMI >25). Those individuals with a BMI
>30 are classified as obese. However, although BMI is widely
used, the amount of body fat is a more important predictor of
mortality and morbidity. Levels of body fat are controlled by
many different factors, but one which has been of great interest
recently is a peptide released by adipocytes, called leptin,
which acts on receptors in the hypothalamus. Genetically obese
mice that lack leptin lose weight when given synthetic leptin.
This discovery was hailed as a cure for obesity, but
unfortunately there are very few obese people who have this
type of genetic mutation. It has now been suggested that obese
people, because they have more fat and secrete more leptin,
have a problem with the sensitivity of their leptin receptors.
C
tissue because it is derived from the same germ layer as the
other connective tissues, the mesoderm, and because it also
consists of cells red blood cells, white blood cells and
platelets in a matrix, the blood plasma. Blood contains the
soluble protein fibrinogen, which is converted into insoluble
fibrin fibres during blood clotting. The different elements of
blood provide metabolic support by carrying nutrients and
waste products and immune protection, by transporting
white blood cells and antibodies around the body. Platelets
are crucial in the control of bleeding, by plugging small holes
in blood vessels and by their involvement in the clotting
cascade (see Ch. 12).
Muscle
Muscle tissue is made up of contractile cells. While there
are some cells in the body that do not form muscle tissues
but are contractile, such as pericytes which surround blood
vessels, most muscle cells (Fig. 2.52) form one of three types
of muscle:

Skeletal muscle
Smooth muscle
Cardiac muscle.

All muscle cells contain the contractile proteins, actin and

myosin, although their arrangement differs between muscle
types. Some classifications separate muscle into:
Voluntary muscles (in that they are under conscious
control, such as the muscles that move elements of the
skeleton)
Involuntary muscles (which are controlled by the
autonomic nervous system and circulating hormones).

In this classification skeletal muscle is classed as volun

tary, and smooth muscle as involuntary (Table 2.28).
Skeletal muscle
This is made up of long, multinucleated cells, called muscle
fibres, which are formed from the fusion of many single cells
during development. Between 10m and 100m in diameter
ch02.indd 85
Fig. 2.52 Muscle types: (A) skeletal muscle; (B) smooth

muscle; (C) cardiac muscle.
and sometimes extending the whole length of the muscle

(i.e. many centimetres), they have a distinct banded appear
ance. This is due to the regular arrangement of the myosin
and actin filaments into repeating units called sarcomeres.
The banded appearance gives rise to the name striated
muscle. The myofilaments which make up the sarcomeres
are arranged in cylindrical myofibrils, which are themselves
grouped together to form the muscle fibres.
The shortening of muscle during contraction occurs due
to overlap between the thin actin and thick myosin filaments,
a process that is stimulated by the release of calcium from
intracellular stores. The stores are called sarcoplasmic
reticulum and form a network of flattened sacs surrounding
the sarcomeres and connected via T-tubules to the surface
of the fibre (see Ch. 9). The contraction of all skeletal muscle
is controlled by the central nervous system via specialised
nerve junctions known as neuromuscular junctions.
Muscle fibres are studded with muscle precursor cells
called satellite cells. These quiescent cells can resume
proliferation when the muscle is either damaged or stressed
in order to replace or increase the muscle mass. This activity
is particularly obvious in weight lifters and body builders, who
can remodel their muscles by increasingly stressing them.
Smooth muscle
Smooth muscle is made up of elongated spindle-shaped
cells that taper at the ends. They usually have a single
nucleus and are much shorter than skeletal muscle cells.
They are often joined together in sheets and the cells are
electrically coupled through gap junctions between adjacent
2/26/2009 6:49:00 PM

Table 2.28 Comparing the different types of muscle
Skeletal
Smooth
Cardiac
Morphology
Multinucleated, long, thin
Single nucleus, spindle-shaped
Single or double nuclei, cylindrical,

branched, prominent intercalated
discs
Appearance
Striated appearance due to

overlapping bands of actin
and myosin
No obvious striations
Striated appearance, but less

organised than skeletal muscle
Unit of excitation
Groups of fibres called

motor units
Linked by gap junctions
Linked by gap junctions
Source of calcium
Calcium stores in extensive

SR
Variable, calcium release can be

induced from rudimentary calcium
stores in SR, calcium can also
enter from extracellular fluid
Calcium enters from extracellular fluid

and releases more calcium from less
extensive SR (calcium-induced
calcium release)
Role of calcium
Calcium removes inhibition
Calcium increases phosphorylation

of contractile proteins
Calcium removes inhibition
Contraction
Discontinuous
Usually continuous (resting tone),

often rhythmic (wavelike), can
maintain very high forces
Continuous, rhythmic
Regenerative ability
Can undergo limited

regeneration (from satellite
cells)
Can regenerate
No regeneration
Activation
Only contracts on nervous

activation by motor nerves
Myogenic but also under

autonomic and hormonal control
Myogenic but also under autonomic

and hormonal control
Receptor/s
Nicotinic cholinergic
Various but include - and

-adrenergic
-adrenergic, muscarinic cholinergic
Neurotransmitter
Acetylcholine
Adrenaline, noradrenaline plus

others
Adrenaline, noradrenaline,
acetylcholine
SR, sarcoplasmic reticulum.
cells to form a functional syncytium. This enables cells to

contract in a coordinated manner with quite diffuse inputs.
Smooth muscle is specialised in producing relatively slow,
low force contractions with a low energy requirement.
However, it can also produce very forceful contractions
under certain circumstances; for example, the contractions
of the uterus during labour.
Muscle contraction is often spontaneous and may be
affected by input from the autonomic nervous system, hor
mones and local factors, such as stretching. In this sense it
is not thought to be under conscious control, hence the term
involuntary muscle, although some smooth muscle can be
controlled consciously.
Smooth muscle cells retain their ability to divide and
hypertrophy in response to increased stress. This is a
problem in high blood pressure, hypertension, where the
response of many blood vessels to the increase in blood
pressure is to increase the thickness of the smooth muscle
layer. This reduces the diameter of the lumen, which in turn
increases the resistance to blood flow and thus further
increases the arterial blood pressure (see Ch. 11).
large numbers of mitochondria and have a rich blood

supply.
Specialised regions of heart muscle are electrically
unstable and form pacemaker areas that trigger rhythmic
muscular contraction in the absence of nervous input. These
electrical signals, and their subsequent contractions, are
then transmitted throughout the heart via low resistance
pathways that ensure the coordinated contraction of the dif
ferent areas of the heart, in order to ensure the pumping of
blood. This intrinsic rhythm is modified by both nervous and
hormonal inputs (see Ch. 11).
There are no stem cells in cardiac muscle, so that, when
heart muscle is damaged, it cannot regenerate and healing
replaces dead muscle with fibrous scar tissue. Heart muscle
can also hypertrophy due to an increase in the size of indi
vidual cells and the addition of myofibrils. Current research
is investigating whether stem cells from other parts of the
body can be injected into heart muscle and be induced to
replace damaged cardiac muscle.
Cardiac muscle
The nervous tissue of the body is divided into:
Cardiac muscle makes up the walls of the heart and is in

some ways intermediate in type between skeletal and smooth
muscle. In appearance, cardiac muscle contains myofibrils
and the muscle appears striated, although the muscle cells
are much shorter than in skeletal muscle. Cardiac muscle
fibres (and myofibrils) are branched and the cells connected
by intercalated discs. These have large numbers of gap
junctions, allowing electrical communication between cells,
and adhering junctions, which bind the cells together to
form a network of electrically and mechanically intercon
nected cells. Due to their high metabolic demands they have
ch02.indd 86
Nervous tissue
The central nervous system (CNS), which consists of
the brain and spinal cord
The peripheral nervous system (PNS), which consists
of all nervous tissue found outside the brain and spinal
cord.

Both the central and peripheral nervous systems are

principally made up of two types of cells: neurons and glia
(see also Ch. 8). There are a few other minor types of cells
also present in the brain: ependymal cells (epithelial cells,
which line the cavities of the brain, or ventricles) and choroid
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Dendritic spine
Dendrites
Cell body
Nucleus
(with nucleolus)
Axon hillock
(trigger zone)
Schwann cell
Axon
Information box 2.22 Neural stem cells
During the development of the nervous system, cells in the

neural tube become multipotent stem cells that can divide
into all the different cells of the brain. Other types of stem
cells in other tissues produce other cells; for example
haematopoietic stem cells in the bone marrow can
produce all the blood cells. While these blood stem cells
remain active throughout life, continually replacing cells,
until recently it was thought that stem cells were not
present in the nervous system after the initial phase of brain
development. However, neural stem cells have now been
isolated from both fetal and adult brains. These cells do not
normally divide in adults, but current research is trying to
identify the exact conditions under which these cells can be
stimulated to divide and grow into mature functioning brain
cells.
Node of Ranvier
Myelin sheath
Synapse
Fig. 2.53 A typical multi-polar neuron.
epithelial cells (which are involved in the secretion of the

cerebrospinal fluid which bathes the brain).
Neurons
Neurons are the cells of the brain that receive signals,
process them and transmit the appropriate response either
to another neuron or to an effector, such as a secretory cell
or a muscle. Neurons come in many different shapes and
sizes but in their most basic form consist of three elements
(Fig. 2.53):
1. A central cell body that contains the cell nucleus and
other intracellular organelles concerned particularly with
protein synthesis and secretory processes. The large
numbers of ribosomes, particularly those associated
with the rough ER or present as polyribosomes, appear
as darkly staining Nissl bodies.
2. Highly branched processes extending from the cell
body called dendrites, which may be covered with
small spines, are the sites of the inputs from other
cells. Dendrites form the main receiving portion of the
neuron.
3. A single axon extending from the cell body to the target
cell. Axons may extend just a few millimetres to a
nearby cell or, as in the case of the motor neurons
supplying muscles distant from the spinal cord, they
may extend for metres. At the start of the axon is an
area called the axon hillock or trigger zone where
electrical signals, known as action potentials, can be
generated. These are then propagated along the axon.
Most neurons can be classified into one of three major
types, depending largely on the position and number of den
drites and the position of the trigger zone. They are:

Multi-polar neurons
Bipolar neurons
Pseudo-unipolar neurons.

ch02.indd 87
Due to their very active nature, neurons in general have

a very high metabolic rate, which means that they need a
continuous secure supply of oxygen and glucose. Because
of this, interruption in the supply is critical, since deprived
neurons will start to die very rapidly. This is particularly prob
lematical as under most conditions mature neurons do not
divide. During development epithelial cells lining the neural
tube give rise to neuroblasts. These cells divide mitotically
to produce amitotic neurons, which then migrate to their final
positions in the brain. As this occurs during fetal develop
ment and is completed in early childhood, the mature brain
does not contain neurons which can divide (however, see
Information box 2.22). Thus neurons that die cannot usually
be replaced. One result of this is that brain tumours derived
from neurons are very rare and occur almost exclusively as
neuroblastomas in children.
Glial cells
The other main category of cell in the nervous system are
glial cells. They make up about half the brain mass and out
number neurons about 10-fold. These cells have a number
of functions, including structural and metabolic support,
immune functions and electrical insulation of axons. There
are three types of glial cells:

Astrocytes
Microglia
Oligodendrocytes and Schwann cells.

Astrocytes
In the CNS the most common type of glial cells are astrocytes. They can be distinguished from neurons by the
presence of a specific protein, glial fibrillary acidic protein
(GFAP). Fibrous astrocytes contain large numbers of fila
ments and are found in the nerve bundles of the white matter
of the brain. Protoplasmic astrocytes have fewer filaments
and are found mainly in the grey matter. They have a number
of fine processes that surround neurons, capillaries and
the ependymal cells lining the ventricles. These astrocytic
endfeet do not touch the capillaries but release factors that
induce bloodbrain barrier characteristics in the capillary
endothelial cells (see Ch. 8). As well as this role, astrocytes
are important in regulating K+ levels around neurons and
providing a store of glycogen that can be supplied to neurons,
in the form of lactate, when required.
Astrocytes surrounding neurons have an important role
in controlling the distribution of neurotransmitter chemicals
2/26/2009 6:49:00 PM

released by the neurons. They do this in two ways, firstly by
restricting the diffusion, and secondly by transporting neuro
transmitters into the astrocyte, where they can be metabo
lised or recycled. The precursors of astrocytes are radial glial
cells which, during early development, span the cerebral
cortex forming a scaffolding for the migration of new nerve
cells to their destinations.
Microglia
Microglial cells are small cells of the CNS with long spiny
processes. During development, in addition to releasing
growth factors, they act as macrophages, removing debris
produced by the programmed cell death which occurs at this
time. In the adult they are normally relatively inactive; however
in the presence of almost any type of injury or insult to the
nervous system, they revert to the role of phagocytotic
macrophages, when they are said to be reactive.
Oligodendrocytes and Schwann cells

Oligodendrocytes and Schwann cells are both involved in
electrically insulating axons in the CNS and PNS, respec
tively. They are large cells with few processes that wrap
around the axons forming multiple lipid bilayers with their
plasma membranes, called a myelin sheath. These mem
branes have a high lipid to protein ratio which makes them
excellent insulators. Oligodendrocytes can myelinate more
than one axon and the cell body lies between them. Schwann
cells only myelinate a single axon and the cell body is closely
apposed to the myelin sheath.
Schwann cells have a role in the regeneration of peri
pheral axons following injury. In order for a peripheral nerve
to re-grow, the tip of the axon must make contact with a
Schwann cell. This stimulates mitosis in the Schwann cell,
which then extends processes towards the growth cone of
the axon. The axon re-grows at between 2mm and 5mm
per day along the Schwann cells, which then re-myelinate
the new axon.
Damaged neurons in the CNS do not seem to regenerate
successfully because CNS glial cells release factors that
specifically inhibit axon growth, although current research
into the supply of neuronal growth factors is encouraging.
The implantation of fetal cells as in the experimental
treatment of Parkinsons disease may prove useful, and
other strategies involving the use of neural stem cells are
ongoing.
Not all axons are myelinated, and this can be seen in
the brain and spinal cord as grey matter, which consists of
cell bodies and unmyelinated axons, as opposed to white
ch02.indd 88
matter, which consists of axons, most of which are myeli

nated. Dendrites are never myelinated.
ORGANS AND SYSTEMS

Organs are made up of at least two tissues, although many
organs contain many more. Organs form distinct structural
units with a particular function/s. Examples of specific organs
are:

Liver
Kidney
Lung
Heart.

Many functions of the body require more than one organ.

For example, the cardiovascular system requires that both
the heart and blood vessels work together in a coordinated
fashion. Some of these divisions may seem somewhat artifi
cial; for example the endocrine system, which consists of
all the organs which secrete hormones, is stimulated by
feedback from almost all the other organs of the body, and
has effects on the entire organism.
Coordinated activity of many

organ systems
At the next level up the whole organism requires the action
of all the organ systems acting together to perform many
functions. For example, the relatively simple activity of
running:

The respiratory system needs to increase its activity to

increase the oxygenation of the blood to meet the
increased oxygen demand.
The cardiovascular system needs to pump more blood
and redirect blood to the appropriate muscles, without
compromising the supply to the brain and the heart.
The nervous system has to initiate and coordinate the
muscular activity, which depends on the locomotor
system for its execution.
The metabolic activities of the liver and other tissues
need to be changed in order to supply the energy
required for the muscle to continue to contract.
When the person stops running, the alimentary system
will absorb the necessary nutrients to replace those
used and the urinary system will excrete waste products
and regulate fluid and acidbase balance.
The immune system will clear away any damaged cells,
and repair mechanisms will rebuild damaged tissues.
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Biochemistry and cell biology introduction

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Biochemistry and cell biology introduction

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Biochemistry and cell biology

Basic cell structure

Protein synthesis and processing

Membrane structure and functions

Specialised plasma membrane structures

Cell division and DNA replication

Key metabolic pathways

Tissues and organs

large molecules from smaller ones, or simply the conversion

Catabolism and anabolism

16 Biochemistry and cell biology

Protozoans, algae, slime moulds

Plasmodium spp. (malaria)

Yeasts, moulds, mushrooms

Candida albicans (candidiasis or thrush)

Mosses, ferns, conifers, flowering plants

None, although many plants are poisonous to humans

Sponges, corals, arthropods, amphibians, reptiles, birds, mammals

Taenia solium (tapeworm)

Table 2.2 Chemical elements of the human body

Abundance (% wet weight)

Minor elements (in order of abundance)

Trace elements (in alphabetical order)

are between 1m and 10m and many eukaryotic cells are

The strength of the different types of bond varies

When dissolved in water many atoms are able to form

Atoms which gain an electron become negatively

These ions will be attracted to each other by the opposite

2.1 Different types of chemical bond.

Table 2.3 Energy of some typical chemical bonds in

Van der Waals interaction

and neutrons (except ordinary hydrogen which has a single

The innermost shell can contain a maximum of two

elements which exist are the noble gases such as helium or

Sodium chloride: an example of an ionic bond

18 Biochemistry and cell biology

Carboncarbon double bonds

On the same side of the double bond (cis)

This isomerism is important in the structures of fatty

Polar and non-polar covalent bonds

Other molecular interactions

Van der Waals interactions

Van der Waals interactions happen when transient

Some reactions are thermodynamically irreversible; that

Oxidationreduction (redox) reactions

couple exergonic and endergonic reactions to transfer energy

INORGANIC MOLECULES AND IONS

glucose + oxygen carbon dioxide + water

The breaking of chemical bonds requires energy input,

Water makes up about 60% of the human body by weight.

Energy in chemical reactions

Either a net release of energy (exergonic reactions)

In exergonic reactions the energy required to break exist

Intracellular fluid contained within cells: about 28L

20 Biochemistry and cell biology

Table 2.4 Common functional groups and classes

Acids, bases and salts

Acids, bases and salts are organic or inorganic compounds

Acids form hydrogen ions (H+) and anions

Bases form hydroxyl ions (OH ) and cations

Acids and bases are crucially important in the control of